[Federal Register Volume 77, Number 86 (Thursday, May 3, 2012)]
[Rules and Regulations]
[Pages 26162-26175]
[FR Doc No: 2012-10649]

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DEPARTMENT OF HEALTH AND HUMAN SERVICES

Food and Drug Administration

21 CFR Parts 600, 610, and 680

[Docket No. FDA-2011-N-0080]

Amendments to Sterility Test Requirements for Biological Products

AGENCY: Food and Drug Administration, HHS.

ACTION: Final rule.

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SUMMARY: The Food and Drug Administration (FDA) is amending the
sterility test requirements for biological products. This rule provides
manufacturers of biological products greater flexibility, as
appropriate, and encourages use of the most appropriate and state-of-
the-art test methods for assuring the safety of biological

products. FDA is taking this action as part of its ongoing efforts to
comprehensively review and, as necessary, revise its regulations
related to biological products.

DATES: This rule is effective June 4, 2012.

FOR FURTHER INFORMATION CONTACT: Paul E. Levine, Jr., Center for
Biologics Evaluation and Research (HFM-17), Food and Drug
Administration, 1401 Rockville Pike, suite 200N, Rockville, MD 20852-
1448,<PHONE_NUMBER>.

SUPPLEMENTARY INFORMATION:

Table of Contents

I. Background
II. Summary of the Final Rule
III. Comments on the Proposed Rule and FDA's Responses
    A. General Comments and FDA's Responses
    B. Comments and FDA's Responses on Specific Topics From the
Proposed Rule
IV. Revisions to Other Regulations
V. Legal Authority
VI. Analysis of Impacts
VII. Environmental Impact
VIII. Federalism
IX. The Paperwork Reduction Act of 1995

I. Background

    This rule revises the sterility requirements for most biological
products under title 21 of the Code of Federal Regulations (CFR),
subchapter F, parts 600 through 680 (21 CFR parts 600 through 680) \1\
and is intended to promote improvement and innovation in the
development of sterility test methods by allowing manufacturers the
flexibility needed for sterility testing of some novel products that
may be introduced to the market, enhancing sterility testing of
currently approved products, and encouraging manufacturers to utilize
scientific and technological advances in sterility test methods as they
become available.
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    \1\ The sterility test provisions of this regulation do not
apply to Whole Blood, Cryoprecipitated Antihemophilic Factor (AHF),
Platelets, Red Blood Cells, Plasma, Source Plasma, Smallpox Vaccine,
Reagent Red Blood Cells, Anti-Human Globulin, or Blood Grouping
Reagents. The provisions also do not apply in cases where the
Director of the Center for Biologics Evaluation and Research (CBER)
or the Director of the Center for Drug Evaluation and Research
(CDER), as appropriate, exempts a product from the requirements
because the Director finds the manufacturer's data adequate to
establish that the mode of administration, the method of
preparation, or the special nature of the product precludes or does
not require a sterility test or that the sterility of the lot is not
necessary to assure the safety, purity, and potency of the product.
(See 21 CFR 610.12(g)(4).)
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    In the Federal Register of June 21, 2011 (76 FR 36019), FDA
published a proposed rule that proposed revisions to update
requirements for sterility testing of biological products. As described
in the preamble of the proposed rule (76 FR 36019 at 36019 to 36020),
any product that purports to be sterile should be free of viable
contaminating microorganisms to assure product safety (Sec.  600.3(q)
(21 CFR 600.3(q)). Absolute sterility of a lot cannot be practically
demonstrated without complete destruction of every finished article in
that lot (USP, Chapter 1211). Therefore, sterility assurance is
accomplished primarily by validation of the sterilization process or of
aseptic processing under current good manufacturing practice (CGMP),
and is supported by sterility testing using validated and verified test
methods (see e.g., USP Chapter 71, European Pharmacopeia 2.6.1.).
    In the Federal Register of November 20, 1973 (38 FR 32048), we
reorganized and republished the biologics regulations, which included
regulations governing sterility testing, as parts 600 through 680.
    Over the years, FDA has amended the biologics regulations, as
necessary, to clarify and update the sterility test requirements. On
March 11, 1976 (41 FR 10427) and March 2, 1979 (44 FR 11754), we
updated Sec.  610.12 (21 CFR 610.12) to clarify the procedures for
repeat testing. On December 15, 1986 (51 FR 44903), we clarified and
updated certain requirements for sterility testing to ensure the
reliability of the growth-promoting qualities of the sterility test
culture media and to provide greater consistency with the test methods
of USP XXI. Finally, on September 15, 1997 (62 FR 48174), we
incorporated by reference into Sec.  610.12(f) the 1995 edition of the
USP concerning the procedures for the membrane filtration test method.
    Prior to this final rule, Sec.  610.12 required that the sterility
of most licensed biological products \2\ be demonstrated through the
performance of tests prescribed in Sec.  610.12(a) and (b).
Specifically, Sec.  610.12 provided that the sterility of each lot of
each product, with the exception of certain products,\3\ be
demonstrated by the performance of prescribed sterility tests for both
bulk and final container material, unless different sterility tests
were prescribed in the license (see Sec.  610.12(g)(1)) or the
manufacturer submitted adequate data \4\ establishing that the mode of
administration, the method of preparation, or the special nature of the
product precluded or did not require a sterility test, or that the
sterility of the lot was not necessary to assure the safety, purity,
and potency of the product (Sec.  610.12(g)(4)(ii)).
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    \2\ See list of exemptions in Sec.  610.12(g)(4).
    \3\ Whole Blood, Cryoprecipitated AHF, Platelets, Red Blood
Cells, Plasma, Source Plasma, Smallpox Vaccine, Reagent Red Blood
Cells, Anti-Human Globulin, or Blood Grouping Reagents (Sec.
610.12(g)(4)(i)).
    \4\ In such an instance, the Director of CBER or CDER, as
appropriate, would determine the adequacy of the data (Sec.
610.12(g)(4)(ii)).
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    The regulation also specified the test method and culture media to
be used. For example, the prescribed sterility test methods relied upon
culture media (either Fluid Thioglycollate Medium or Soybean-Casein
Digest Medium) to detect growth of microorganisms (Sec.  610.12(a)(1)
and (a)(2)). Moreover, Sec.  610.12 specified criteria, such as
incubation conditions (time and temperature) to be used during testing,
suitable test organisms for the evaluation of the growth-promoting
qualities of the culture media, storage and maintenance of test
organism cultures, and storage and condition of media.
    Since we last clarified and updated our regulations governing
sterility testing, advances in technology in recent years have allowed
the development of new sterility test methods that yield accurate and
reliable test results in less time and with less operator intervention
than the currently prescribed methods. Some examples of novel methods
include the Adenosine Triphosphate bioluminescence, chemiluminescence,
and carbon dioxide head space measurement. Manufacturers may benefit
from using such sterility test methods with rapid and advanced
detection capabilities.
    Accordingly, we have amended Sec.  610.12 to promote improvement
and innovation in the development of sterility test methods, to address
the challenges of novel products that may be introduced to the market
in the future, and to potentially enhance sterility testing of
currently approved products. This final rule provides manufacturers the
flexibility to take advantage of methods as they become available,
provided that these methods meet certain criteria.

II. Summary of the Final Rule

    FDA is adopting as final, without material change, the proposed
requirements for sterility testing. Specifically, this final rule:
     Eliminates specified sterility test methods, culture media
formulae (or formulation), and culture media test requirements;
     Eliminates specified membrane filtration procedure
requirements for certain products;
     Eliminates specified sterility test requirements for most
bulk material;

     Modifies the repeat sterility test requirements, so that
repeat tests will occur only once for each lot. These repeat tests are
limited to situations when the quality control unit conclusively
determines, after conducting an investigation upon detection of viable
microbial contamination during the initial test of the lot, that the
contamination is the result of laboratory error or faulty materials
used in conducting the sterility test;
     Replaces the storage and maintenance requirements for
cultures of test organisms used to determine the ``growth-promoting
qualities'' of culture media with: (1) Validation requirements
specifying that any sterility test used is able to consistently detect
the presence of viable contaminating microorganisms and (2)
verification of ``growth-promoting properties'' or microorganism-
detection capabilities of test and test components;
     Replaces the sample size or amount requirement with a
requirement that the sample be appropriate to the material being
tested;
     Replaces the Interpretation of test results section under
Sec.  610.12(c) with a requirement that manufacturers establish,
implement, and follow written procedures for sterility testing that
describe, at a minimum, the test method used, the method of sampling,
and the written specifications for acceptance or rejection of each lot;
     Simplifies and clarifies the Exceptions section under
Sec.  610.12(h); and
     Identifies the Director of CDER as one of the two Center
directors authorized to grant an exemption under the exception
provision at Sec.  610.12(h)(2). In the proposed rule, the Center for
Devices and Radiological Health was erroneously identified in this
exception, instead of the Center for Drug Evaluation and Research.
     Revises the definition of the term ``sterility'' under
Sec.  600.3(q); and
     Eliminates certain exceptions for allergenic products
related to sterility testing under Sec.  680.3(c).

III. Comments on the Proposed Rule and FDA's Responses

    We received 17 letters of comments on the proposed rule. These
comments were received from biologics manufacturers, industry
associations, and other interested persons. A summary of the comments
received and our responses follow. We first respond to general comments
and then respond to comments on the specific topics set forth in the
preamble of the proposed rule.
    To make it easier to identify the comments and our responses, the
word ``Comment,'' in parentheses, will appear before the comment's
description, and the word ``Response,'' in parentheses, will appear
before our response. We have also numbered each comment to help
distinguish between different comments. The number assigned to each
comment is purely for organizational purposes and does not signify the
comment's value or importance or the order in which it was received.
Certain comments were grouped together because the subject matter of
the comments was similar.

A. General Comments and FDA's Response

    (Comment 1) Thirteen of the letters of comments supported the
proposed rule. Many of the comments agreed that the proposed amendments
would provide manufacturers of biological products greater flexibility
and would promote improvement and innovation in the development of
sterility test methods. Several comments agreed that the proposed
amendments would allow manufacturers to use the most appropriate and
state-of-the-art test methods for assuring the safety of biological
products. Several comments applauded FDA's effort to amend sterility
test requirements to permit the use of new methods and systems in
assessing microbiological contamination in sterile products. Another
comment was pleased to see FDA's commitment to advancing the principles
of innovation in product development for public health.
    (Response) FDA acknowledges and appreciates the supportive
comments. As stated previously, the rule provides needed flexibility
and encourages manufacturers to benefit from scientific and
technological advances in sterility test methods as they become
available.
    (Comment 2) One comment noted an error in the reference to the
European Pharmacopeia 2.6.2. provided in the first paragraph in section
I of the preamble to the proposed rule. The comment pointed out that
European Pharmacopeia 2.6.2. is the chapter for Mycobacteria testing.
    (Response) We agree with this comment. The reference should have
been to European Pharmacopeia 2.6.1. Sterility testing.
    (Comment 3) One comment concurred with the preamble statement that
``* * * sterility assurance is accomplished primarily by validation of
the sterilization process or by the aseptic processing procedures under
CGMP, and is supported by sterility testing using validated and
verified test methods,'' (76 FR 36019 at 36019). However, the commenter
went on to state that ``* * * the regulations would be better suited by
ensuring that the aseptic manufacturing processes follow strict GMP,
further leveraging the requirements for aseptic environments, media
fill programs, and strict oversight of the aseptic process as opposed
to the perceived assurance that sterility testing of samples provides.
This is best illustrated through existing verbiage in Sec.  211.113(b)
(21 CFR 211.113(b)) but should be further expanded upon to provide
improved guidance to industry and investigators.''
    (Response) We acknowledge that product sterility testing does not
provide absolute assurance of product sterility. However, we believe
validation of aseptic processes,\5\ using process simulations or media
fills, together with operational controls and product sterility
testing, provide a sufficient level of assurance that products
purported to be sterile are in fact sterile. Therefore, we do not agree
that additional requirements are necessary because the existing CGMP
requirements under parts 210 and 211 (21 CFR parts 210 and 211) and the
other applicable regulations in parts 600 through 680 already address
the concerns raised by the commenter. We believe this final rule,
together with the other applicable regulations and Agency guidance,
provide manufacturers appropriate latitude to determine how to achieve
the level of control necessary for compliance.
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    \5\ See the applicable requirements in parts 210, 211, and 600
through 680, and FDA's guidance document entitled ``Guidance for
Industry: Sterile Drug Products Produced by Aseptic Processing--
Current Good Manufacturing Practice,'' dated September 2004.
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    (Comment 4) One comment expressed a concern that an environmental
requirement is not part of the proposed rule. The commenter stated,
``Environmental conditions are important to avoid cross-contamination''
and proposed the addition of the following wording described in
European Pharmacopeia 2.6.1. ``The test for sterility is carried out
under aseptic conditions. In order to achieve such conditions, the test
environment has to be adapted to the way in which the sterility test is
performed. The precautions taken to avoid contamination are such that
they do not affect any microorganisms which are to be revealed in the
test. The working conditions in which the tests are performed are
monitored regularly by appropriate sampling of the working area and by
carrying out appropriate controls.''

    (Response) In discussing ``environmental conditions,'' we
understand the comment to mean environmental controls. We have
considered the issue, including the points raised in this comment and
have decided not to adopt the suggested language or revise the rule in
light of the suggested language because the concerns expressed by the
commenter are currently addressed in the CGMP requirements in parts 210
and 211 and the applicable regulations in parts 600 through 680. In
addition, manufacturers may turn to relevant Agency guidance documents
for additional guidance. Furthermore, as the commenter states, the
proposed wording regarding environmental controls under which the
sterility test is to be performed is already described in European
Pharmacopeia 2.6.1., and USP Chapter 71, both of which are additional,
valuable resources for manufacturers.
    (Comment 5) One comment noted that while Sec.  610.12 addresses
aspects of sterility, the current theme of the section is specific to
sterility testing. The commenter therefore suggested either renaming
the title of Sec.  610.12 as ``Sterility Test,'' or broadening Sec.
610.12 so that the regulation addresses all critical elements in the
content area of sterility.
    (Response) We decline to adopt either recommended change because we
believe that the current title of Sec.  610.12 remains appropriate and
that the suggested title change is unnecessary. In response to the
comment expressing a desire to broaden Sec.  610.12 to address all
critical elements in the content area of sterility, FDA notes that this
comment is outside the scope of this final rule.

B. Comments and FDA's Response on Specific Topics From the Proposed
Rule

    The following are comments and FDA's responses, as identified by
the specific topic in the proposed rule to which the comment and FDA's
response applies.
1. When is sterility testing required?
    For the reasons discussed in the preamble to the proposed rule (76
FR 36019 at 36020 to 36021), we proposed amending Sec.  610.12 to
eliminate the sterility test requirement for most bulk materials. We
have determined that, in most cases, for purposes of sterility testing,
the most appropriate test material is the final container material. We
recognize that due to the nature of some biological products, testing
the final container material may not always be feasible or appropriate.
Thus, as finalized, Sec.  610.12 requires that prior to release,
manufacturers of biological products must perform sterility testing of
each lot of each biological product's final container material or other
material (e.g., bulk material or active pharmaceutical ingredient
(API), in-process material, stock concentrate material), as
appropriate, and as approved in the biologics license application (BLA)
or BLA supplement. For example, as discussed in the preamble to the
proposed rule (76 FR 36019 at 36021), certain allergenic and cell and
gene therapy products may need to be tested for sterility at an in-
process stage or some other stage of the manufacturing process (e.g.,
intermediate, API, bulk drug substance) instead of the final container
material because the final container material may interfere with the
sterility test. Likewise, as discussed in the preamble to the proposed
rule, some cell therapy products and cell-based gene therapy products
may need to be tested for sterility at an in-process stage or some
other stage of manufacturing process because low production volumes may
result in an insufficient final container material sample for sterility
testing or a short product shelf-life may necessitate administration of
the final product to a patient before sterility test results on the
final container material are available.
    (Comment 6) Three comments were particularly supportive of FDA's
proposal to eliminate the sterility test requirements for bulk
material. One comment noted this change will be particularly helpful
for cellular therapy products.
    (Response) We appreciate the supportive comments. We agree that the
elimination of specified sterility test requirements for most bulk
materials will provide manufacturers with greater flexibility and in
most cases, for purposes of sterility testing, the most appropriate
test material is the final container (76 FR 36019 at 36021). We also
acknowledge that due to the nature of some biological products, this
change could result in the need for some manufacturers to modify their
testing procedures to eliminate testing for bulk materials. However, we
note that these modifications to eliminate testing for bulk materials
would be made following existing change control procedures and a
submission to FDA to report the change would not be required.
    If it is determined that sterility testing needs to be performed on
material other than the final product, due to the nature of the final
product, we would expect the manufacturer, as required under Sec. Sec.
601.2 and 601.12, to include in its BLA or BLA supplement: (1) A
description of the details of the sterility test method used, including
the procedure for testing the alternate material instead of the final
container material; and (2) the scientific rationale for selecting the
specific test material instead of the final container material.
    As discussed in the preamble to the proposed rule (76 FR 36019 at
36021), a manufacturer who desires to utilize an alternate sterility
test method other than the one approved in its BLA must submit a BLA
supplement in accordance with Sec.  601.12(b).
    (Comment 7) One comment asserted that upon finalization of the
rule, a manufacturer who desires to utilize an alternative sterility
test other than the one approved in its BLA should be permitted to
submit the change to FDA in its annual report in accordance with Sec.
601.12(d), as opposed to a prior approval supplement to an approved
application under Sec.  601.12(b).
    (Response) We consider changes that may affect the sterility
assurance level of a product to have substantial potential to affect
the safety, purity, or potency of a product and have consistently
identified this change as one that requires prior approval. Therefore,
a manufacturer who desires to utilize an alternate sterility test
method other than the one approved in its BLA must submit a prior
approval supplement to an approved application in accordance with Sec.
601.12(b). We note that approval of the supplement will be based on the
determination that the data submitted with the request establishes a
regulatory basis for approval.
2. What are the sterility test requirements?
    a. Test methods--We proposed amending Sec.  610.12 to eliminate
references to specific test methods and culture media for sterility
testing and to instead require that the sterility test be appropriate
to the material being tested such that the material does not interfere
with or otherwise hinder the test. As discussed in the preamble to the
proposed rule (76 FR 36019 at 36021), we believe this revision
recognizes current practices and provides manufacturers the flexibility
to take advantage of suitable modern sterility test methods and keep
pace with advances in science and technology.
    As also discussed in the preamble to the proposed rule (76 FR 36019
at 36021), because we are expanding potentially acceptable sterility
test methods to include non-culture-based methods in addition to
culture-based methods, we also have removed the definition of ``a lot
of culture medium.'' Previously, Sec.  610.12(e)(2)(i) defined this
term as ``* * * that quantity of uniform material identified as having
been

thoroughly mixed in a single vessel, dispensed into a group of vessels
of the same composition and design, sterilized in a single autoclave
run, and identified in a manner to distinguish one lot from another.''
Although we have deleted this term from Sec.  610.12, we believe (as
stated in the preamble to the proposed rule) that this concept is
captured by the definition of ``lot'' in Sec.  600.3(x). We note that
this change is also consistent with our understanding that prepared
culture media may be purchased, in which case a lot may be
predetermined by the vendor.
    (Comment 8) Two comments opposed the elimination of the specified
sterility test methods and culture media because eliminating the
specific requirements may lead to different interpretations by
industry, as well as FDA investigators. One comment stated that the
current text on acceptable culture media, reference organisms, and
incubation temperatures for sterility testing represents essential
guidance for industry. The comments suggested that either the current
regulations be retained in addition to the proposed amendments or
retained as guidance.
    (Response) We reiterate that the purpose of this rule is to provide
manufacturers of biological products greater flexibility and to
encourage use of the most appropriate and state-of-the-art test methods
for assuring the safety of biological products. Accordingly, at this
time, we decline to retain the current specified sterility test
methods, culture media, reference organisms, and incubation
temperatures in regulation or guidance. Furthermore, we disagree that
this rule may lead to inconsistent interpretations by industry and FDA
staff because sterility test methods for biological products are
approved in the manufacturer's BLA or BLA supplement, and hence, the
data submitted with the request are reviewed in a consistent manner in
accordance with review management procedures. Therefore, we believe the
commenters' concerns about inconsistencies in interpretation are
unfounded.
    (Comment 9) One commenter expressed concern about the applicability
of the proposed changes in the global regulatory market in that the use
of approved alternative sterility methods would not be globally
applicable in the absence of compendial harmonization. The commenter
inquired whether FDA has plans to harmonize the use of alternative
sterility methods with the three main global compendia.
    (Response) We do not agree that the final rule and the use of a
suitable modern sterility test method will interfere with the global
regulatory market. The purpose of the rule is to provide for greater
flexibility and to encourage use of the most appropriate and state-of-
the-art test methods for assuring the safety of biological products. We
believe this final rule will foster the adoption of novel methods and
that alignment with global pharmacopeial methods will occur over time.
With respect to FDA's future plans to harmonize the use of alternative
sterility methods with the three main global compendia, we note that
any such discussion is outside the scope of this rule.
    (Comment 10) One comment proposed adding a reference in the
regulations to a compendial method and allowing for the implementation
of alternative methods. The commenter expressed concern that, in the
global marketplace, implementation of a novel method different from USP
Chapter 71 would not be harmonized with other compendia and might pose
risks to approval of marketing authorizations if new tests are not
recognized or accepted by foreign health authorities.
    (Response) We do not agree with the comment and note that
incorporating such a reference would be inconsistent with the intent of
this rule. We reiterate that we do not agree that this final rule will
interfere with the global marketplace. Rather, we believe that
facilitating flexibility and encouraging the use of the most
appropriate and state-of-the-art test methods will foster the adoption
of novel method technologies and that alignment with pharmacopeia
methods will occur over time. Furthermore, as we have explained in the
preamble to the proposed rule, FDA considers established USP compendial
sterility test methods to already have been validated using an
established validation protocol; therefore their accuracy, specificity,
and reproducibility need not be reestablished to fulfill the validation
requirements under the final rule. Only a manufacturer who desires to
utilize an alternative method other than the one approved in its BLA
must submit a BLA supplement in accordance with Sec.  601.12(b). This
rule does not require manufacturers to utilize an alternative method
other than the one approved in their BLA.
    (Comment 11) One comment stated that the absence of references to
standards such as USP Chapter 71 within Sec.  610.12 may lead to
confusion and suggested that a general disclaimer that FDA is not
endorsing any particular standard or the provision of specific examples
within the regulation may provide an important point of reference for
compliance. Two comments stated that USP Chapter 71 and European
Pharmacopeia 2.6.1. should be listed within Sec.  610.12 as a baseline
or standard for sterility testing. Two other comments recommended
referring to the USP Chapter 71 as the ``referee'' method instead of
referring to it as an example.
    (Response) The concerns expressed in the comments are unfounded. We
reiterate that we consider the current sterility test methods in a
manufacturer's BLA or BLA supplement to already have been validated. In
contrast, newer methods (for example, non-culture-based methods that
have not been validated according to an established protocol) or those
that deviate from the official compendial sterility test methods will
require validation.
    Moreover, the final rule requires that a novel method be validated
in accordance with an established protocol to demonstrate that the test
is capable of consistently detecting the presence of viable
microorganisms. We believe methods validation is a well recognized
activity and can be performed without comparison to a ``referee'' test
method.
    Furthermore, we note that there is no single ``referee'' test
method that would work for all products and that some novel methods
cannot be easily compared to culture-based methods such as USP Chapter
71 because these testing methods do not measure microbial growth.
Therefore, we believe that it is neither necessary nor appropriate to
add a reference to a standard or ``baseline'' in this final rule.
    (Comment 12) We received two comments regarding growth-promotion
testing. One comment asserted that the proposal to eliminate the
requirements to test culture media with specific test organisms, to
eliminate the number of organisms that must be used to demonstrate
growth-promoting qualities of culture media, and to eliminate specific
incubation conditions and visual examination requirements may lead to
different interpretations on which organisms can and should be used.
The comment proposed that a reference to a ``referee'' method be added
to the regulation including requirements for growth promotion and the
strains and number of organisms to be used. The other comment supported
the elimination of the list of specified organisms, while also stating
that providing a list of organisms for manufacturers to consider would
be a benefit to facilities that do not have the necessary expertise or
staffing.
    (Response) Because we are providing manufacturers the flexibility
to use

sterility test methods that are either culture-based or non-culture-
based, which may necessitate different verification activities, we
decline to retain the existing requirements for specified sterility
test reference organisms. For similar reasons, we do not believe a
reference to a ``referee'' method is necessary or appropriate and we
decline to adopt the recommended change.
    Instead of specifying the number and type of test organisms, under
Sec.  610.12(b) of the final rule, we require that: (1) The sterility
test must be appropriate to the material being tested such that the
material does not interfere with or otherwise hinder the test; (2) the
sterility test must be validated to demonstrate that the test is
capable of reliably and consistently detecting the presence of viable
contaminating microorganisms; and (3) the sterility test and test
components must be verified to demonstrate that the test method can
consistently detect the presence of viable contaminating
microorganisms.
    Due to the variety of currently available and potential future
sterility test methods, we have eliminated specified incubation
conditions (time and temperature) and visual examination requirements
previously prescribed in Sec.  610.12. Since we are allowing any
validated sterility test method that is appropriate to the material
being tested, rather than specifying the test and the media used, we
have also eliminated the Fluid Thioglycollate Medium incubation
temperatures previously prescribed in Sec.  610.12(a)(1)(ii) for the
final container material containing a mercurial preservative.
    (Comment 13) One comment recommended that, with respect to
validation, a definition for the terms ``reliably'' and
``consistently'' be added to the regulation for greater utility in
understanding expectations when validating a method. The commenter
offered, for example, ``* * * that a validated method, though
performing consistently and reliably, may still not be centered on the
true value of the specific parameter being tested. Consequently, when
this method would be used during testing the results may be in a
statistical state of control, but not necessarily statistically capable
of measuring the true value.'' The commenter asked FDA to consider ``*
* * that the use of the terms `reliably and consistently' may infer
that the validation of a test for non-sterility does not require proof
of performance at least equivalent to the USP referee method.'' The
comment therefore asked that Sec.  610.12(b)(2) be revised to require
that the sterility test be validated to demonstrate an equivalent or
superior detection of viable contaminating microorganisms compared to
the USP compendial or like method.
    (Response) FDA has considered the issues raised by these comments
and has determined that making the suggested changes would be
inconsistent with the intent of this rule. With respect to the comment
that the rule should be revised to require that the sterility test be
validated to demonstrate an equivalent or superior detection of viable
contaminating microorganisms compared to the USP compendial or like
method, we reiterate that some novel methods cannot be easily compared
to culture-based methods such as USP Chapter 71 because they do not
measure microbial growth. Moreover, we note that the final rule
requires that a novel method be validated in accordance with an
established protocol to demonstrate that the test is capable of
consistently detecting the presence of viable microorganisms. With
respect to the comment that the terms ``reliably'' and ``consistently''
should be defined, we note that these terms are already well understood
in the industry.
    b. Validation--As discussed in the preamble to the proposed rule
(76 FR 36019 at 36021 to 36022), the International Conference on
Harmonisation (ICH) publication entitled ``Validation of Analytical
Procedures: Text and Methodology Q2(R1)'' dated November 2005, states
that ``The objective of validation of an analytical procedure is to
demonstrate that it is suitable for its intended purpose.'' \6\
Similarly, USP General Chapter 1223, ``Validation of Alternative
Microbiological Methods,'' states ``Validation of a microbiological
method is the process by which it is experimentally established that
the performance characteristics of the method meet the requirements for
the intended application.'' For sterility testing, this means that the
test can consistently detect the presence of viable contaminating
microorganisms.
---------------------------------------------------------------------------

    \6\ This guideline for industry was previously named ``Text on
Validation of Analytical Procedures'' (ICH-Q2A), dated March 1995
(approved by the Steering Committee in October 1994). An
accompanying guideline entitled ``Validation of Analytical
Procedures: Methodology (Q2B),'' dated November 6, 1996, was
subsequently developed and approved by the Steering Committee in
November 1996. The parent guideline is now renamed ``Validation of
Analytical Procedures: Text and Methodology Q2(R1)'' and was revised
in November 2005. At that time, the guideline on methodology (Q2B)
was incorporated into the parent guideline.
---------------------------------------------------------------------------

    We have eliminated the prescribed sterility test methods found in
Sec.  610.12 and instead will allow the use of sterility test methods
that are validated in accordance with established protocols to be
capable of consistently detecting the presence of viable contaminating
microorganisms. If an established USP compendial sterility test method
is used, a manufacturer must verify that this established method is
suitable for application to the specific product (see Sec. Sec.
211.165(e) and 211.194(a)); however, FDA considers established USP
compendial sterility test methods to already have been validated using
an established validation protocol, so their accuracy, specificity, and
reproducibility need not be reestablished to fulfill the validation
requirement under the final rule. In contrast, novel methods and any
methods that deviate from the USP compendial sterility test methods
require the detailed validation discussed in this document and
elsewhere in this preamble.
    We again note that Sec.  610.12 requires the use of a material
sample that does not interfere with or otherwise hinder the sterility
test from detecting viable contaminating microorganisms. This
requirement is crucial because the material itself or substances added
to the material during formulation may make some sterility tests
inappropriate for use. A validated sterility test method is a critical
element in assuring the safety, purity, and potency of the product. USP
General Chapter 1223, as well as the ICH guideline referenced earlier
entitled ``Text on Validation of Analytical Procedures,'' dated March
1995 (ICH-Q2A), provide general descriptions of typical validation
parameters, how they are determined, and which subset of each parameter
is required to demonstrate validity, based on the method's intended
use. Validation of each test method should be performed on a case-by-
case basis to ensure that the parameters are appropriate for the
method's intended use. In the context of reviewing sterility test
methods as part of BLAs and BLA supplements, FDA may decide, as
appropriate, to encourage the use of the compendial method as a
benchmark or starting point for validation of novel methods and certain
other methods.
    (Comment 14) One comment requested clarification regarding
validation of novel methods and any methods that deviate from the USP.
This commenter stated that to validate novel test methods, ``the
sponsor not only has to test the matrix effects'', but also has to
validate the new method against the USP compendial method. The

commenter also stated that this would impede the use of innovative
technologies and increase the risk and cost to the sponsor. In
addition, the commenter recommended that duplicative testing
requirements be avoided and that the manufacturer of the technology or
a third party be allowed to perform the validation of new methods.
    (Response) The commenter misinterpreted the validation requirements
under the proposed (and final) rule. The revisions we are adopting in
the final rule do not require duplicative validation of novel methods
against the USP compendial method or testing under a separate
validation procedure. Instead, novel methods and any methods that
deviate from the USP compendial sterility test methods will require a
single, detailed validation study to be conducted, which may include
the use of the compendial method as a benchmark or starting point. We
disagree that such validation will impede the use of innovative
technologies and will increase the risk and cost to the sponsor.
Instead, we believe that, as discussed elsewhere in this document and
in the preamble to the proposed rule, that this final rule will
encourage the use of innovative technology.
    (Comment 15) One comment referenced the preamble statement that ``*
* * FDA may decide, as appropriate, to encourage the use of the
compendial method as a benchmark or starting point for validation of
novel methods and certain other methods.'' (76 FR 36019 at 36022) and
suggested that the use of the compendial method as a benchmark or
starting point should be more strongly encouraged.
    (Response) While FDA may decide, as appropriate, to encourage the
use of the compendial method as a benchmark or starting point for
validation of some novel or other methods, we also may decide not to
encourage such use for some (for example, non-culture-based) methods
that cannot easily be compared to culture-based methods such as the USP
compendial method. Therefore, we disagree that the use of the
compendial method as a benchmark or starting point should be more
strongly encouraged or required.
    (Comment 16) We received two comments in response to our request in
the proposed rule for comments on whether the proposed requirements are
sufficient to ensure adequate validation of novel sterility test
methods or whether additional criteria or guidance is needed. One
comment recommended that any guidance to accompany the final rule be
developed to include such things as a list of organisms for
manufacturers to consider in the development of their validation and
verification plans, including examples of when verification is
required. One comment suggested that such additional guidance include
information related to a determination of the panel of relevant
organisms in the sample matrix used in challenging the sterility test
during validation.
    (Response) We appreciate the interest in additional guidance for
validation of novel sterility test methods and will consider the need
to develop future guidance in accordance with the good guidance
practices set out in 21 CFR 10.115.
    As discussed in the preamble to the proposed rule, it is important
to consider validation principles, such as limit of detection,
specificity, ruggedness, and robustness, while developing the
validation protocol and performing validation studies. These terms are
defined as follows:
     The ``limit of detection'' reflects the lowest number of
microorganisms that can be detected by the method in a sample matrix.
This is necessary to define what is considered contaminated.
     ``Specificity'' is the ability of the test method to
detect a range of organisms necessary for the method to be suitable for
its intended use. This is demonstrated by challenging the sterility
test with a panel of relevant organisms in the sample matrix.
     ``Ruggedness'' is the degree of reproducibility of results
obtained by analysis of the same sample under a variety of normal test
conditions, such as different analysts, different instruments, and
different reagent lots.
     ``Robustness'' is the capacity of the test method to
remain unaffected by small, but deliberate, variations in method
parameters, such as changes in reagent concentration or incubation
temperatures.
    (Comment 17) One comment stated that for the detailed validation of
a novel method, the validation principles should be restricted to the
limit of detection, specificity, and robustness (i.e., to not include
ruggedness).
    (Response) We agree that the validation principles of limit of
detection, specificity, and robustness are important to consider when
developing protocols and performing validation studies. However, we
understand the comment to suggest excluding ruggedness. We view
ruggedness as an important validation principle to be considered, and
we do not agree with excluding it from the scope of this rule. We note
that the final rule does not include prescriptive details on how to
conduct validation studies; it simply codifies our longstanding policy
that the sterility test must be validated to demonstrate that the test
is capable of reliably and consistently detecting the presence of
viable contaminating microorganisms.
    (Comment 18) One comment objected to the requirement in existing
Sec.  211.160(b) as to the establishment of sampling plans because ``*
* * it is not practical or feasible to develop a scientifically sound
sampling plan to ensure a product conforms to standards of sterility.''
The comment recommended as a solution to either remove the requirement
for scientific sampling plans with respect to sterility testing or to
provide a clarification of ``scientifically sound'' versus
``appropriate.''
    (Response) The suggested revisions go beyond the scope of the
proposed changes to the sterility test requirements. Furthermore, Sec.
211.160(b) is an existing current good manufacturing practice
requirement for finished pharmaceuticals, which states that laboratory
controls must include the establishment of scientifically sound and
appropriate specifications, standards, sampling plans, and test
procedures designed to assure that components, drug product containers,
closures, in-process materials, labeling, and drug products conform to
appropriate standards of identity, strength, quality, and purity. We
consider such laboratory controls to be needed for both culture-based
and non-culture-based sterility test methods. As stated in the preamble
to the proposed rule (76 FR 36019 at 36022), the manufacturer must
establish and document the test method's accuracy, sensitivity,
specificity, and reproducibility (Sec.  211.165(e)), as specified in
the BLA or BLA supplement (Sec. Sec.  601.2, 601.12). For sterility
tests, FDA believes that a validation protocol that would meet these
standards would, at a minimum, include samples of the material to be
marketed and incorporate appropriate viable contaminating
microorganisms to demonstrate the sterility test's growth-promoting
properties or the method's detection system capabilities, depending on
the type of test method used. In addition, validation protocols for
culture-based methods should include both aerobic and anaerobic
microorganisms when selecting test organisms and include microorganisms
that grow at differing rates so that manufacturers can establish that
the test media are capable of supporting the growth of a wide range of
microorganisms.

    When utilizing culture-based methods, where appropriate, validation
protocols should require that challenge organisms be added directly to
the product prior to membrane filtration or direct inoculation. If this
is not possible due to inhibition by the product, then validation
protocols should require that the challenge organism be added to the
final portion of sterile diluent used to rinse the filter, if a
membrane filtration test method is used, or directly to the media
containing the product if a direct inoculation test method is used.
    For non-culture-based methods, the feasibility of identifying
microorganisms from a contaminated sample should be evaluated during
validation. If a method does not have the capability to identify
microorganisms to the species level, the validation protocol should
require that an additional method for species identification be
utilized for investigation of detected contaminants. The test organisms
selected should reflect organisms that could be found in the product,
process, or manufacturing environment.
    (Comment 19) Two comments sought clarification of the following
statement in the preamble to the proposed rule: ``When utilizing
culture-based methods, validation protocols should require that
challenge organisms be added directly to the product prior to membrane
filtration or direct inoculation. If this is not possible due to
inhibition by the product, then validation protocols should require
that the challenge organism be added to the final portion of sterile
diluent used to rinse the filter if a membrane filtration test method
is used, or directly to the media containing the product if a direct
inoculation test method is used.'' (76 FR 36019 at 36022)
    One commenter stated that this language is inconsistent with the
harmonized compendial method suitability test which states, ``After
transferring the content of a container or containers to be tested to
the membrane, add an inoculum of small number of viable microorganisms
(not more that 100 colony-forming units) to the final portion of
sterile diluents used to rinse the filter.'' Another comment sought
clarification of the suggested limits for the density of the inoculum
of challenge organisms added directly to the product.
    (Response) The intent of these statements was to clarify that for
certain biological products utilizing culture-based methods, method
suitability testing necessitates adding the challenge organism directly
to the product prior to membrane filtration or direct inoculation.
Therefore, we are now clarifying that when utilizing culture-based
methods, where appropriate, validation protocols should require that
challenge organisms be added directly to the product before membrane
filtration or direct inoculation. If this is not possible due to
inhibition by the product, then validation protocols should require
that the challenge organism be added to the final portion of sterile
diluent used to rinse the filter if a membrane filtration test method
is used or directly to the media containing the product if a direct
inoculation test method is used.
    (Comment 20) One comment addressed the selection of organisms to be
used. The comment suggested that with respect to validation protocols,
for consistency, the wording regarding the selection of organisms
should specifically include wild-type isolates that have been recovered
from the controlled manufacturing environment and past contaminants of
the product or any of its sterile components. The comment also
suggested that this requirement should extend beyond culture-based
methods. Further, the comment suggested that the statement in the
preamble that `` `The test organisms selected should reflect organisms
that could be found in the product, process, or manufacturing
environment (emphasis added) [76 FR 36019 at 36022],' should be
tightened to require use of strains actually isolated from the product,
process, or manufacturing environment, as the word `reflect' probably
implies use of relevant species that might be sourced from culture
collections rather than explicitly requiring use of wild-type strains
(plant isolates).''
    (Response) Our intention with respect to this statement was to
include those organisms recovered both from the controlled
manufacturing environment and from the product. Furthermore, the
preamble statement was intended to refer to validation protocols in
general, where appropriate, to both culture-based and non-culture-based
test methods.
    The validation study design should contain the appropriate controls
to evaluate the product sample's potential to generate false-positive
and false-negative results. Validation of the sterility test should be
performed on all new products, and repeated whenever there are changes
in the test method or production method that could potentially inhibit
or enhance detection of viable contaminating microorganisms.
    (Comment 21) One comment recommended the addition of ``or
production method'' to the statement in the preamble so that it would
now read, ``Validation of the sterility test should be performed on all
new products, and repeated whenever there are changes in the test
method or production method that could potentially inhibit or enhance
detection of viable contaminating microorganisms.'' (See original
statement 76 FR 36019 at 36022.) The commenter stated that the
additional language is appropriate because the production process may
influence the matrix of the test article, which may in turn influence
the sterility test verification.
    (Response) We agree that changes in the production method or
manufacturing process could affect the results of testing conducted on
the product. Therefore, we agree that validation of the sterility test
should be performed on all new products and repeated whenever there are
changes in the test method or production method that could potentially
inhibit or enhance detection of viable contaminating microorganisms.
    c. Verification--As stated in the proposed rule (76 FR 36019 at
36022), verification is the confirmation that specified requirements
have been fulfilled as determined by examination and provision of
objective evidence. While validation of a sterility test method is the
initial process of demonstrating that the procedure is suitable to
detect viable contaminating microorganisms, verification occurs over
the lifetime of the sterility test method and is the process of
confirming that the sterility test and test components continue to be
capable of consistently detecting viable contaminating microorganisms
in the samples analyzed. This verification activity may be necessary on
a periodic basis or each time a sample is tested, depending upon the
test method used. Under Sec.  610.12(e) of the final rule, we require
that the sterility test and test components be verified, as
appropriate, to demonstrate that they can continue to consistently
detect viable contaminating microorganisms.
    (Comment 22) One comment maintained that the section of the
preamble to the proposed rule regarding verification was not totally
clear and should be reworded to explain the intended purpose.
Specifically, the comment suggested, in order to clarify the goal of
verification, adding the following sentence, ``The intended purpose of
the verification is to confirm that all the reagents utilized in the
sterility test are qualified.'' The commenter also noted that
validation is to be done using the product to be tested and proposed
adding the phrase ``in the product to be tested'' to the following
statement in the preamble ``While

validation of a sterility test method is the initial process of
demonstrating that the procedure is suitable to detect viable
contaminating microorganisms, verification occurs over the lifetime of
the sterility test method and is the process of confirming that the
sterility test and test components continue to be capable of
consistently detecting viable contaminating microorganisms in the
samples analyzed.'' (76 FR 36019 at 36022 to 36023)
    (Response) To the extent that the commenter is arguing that our
explanation is unclear, we disagree. As stated in the preamble to the
proposed rule at section III.E (76 FR 36019 at 36022 to 36023), we
believe that in order to verify the sterility test, verification
activities are necessary to demonstrate that sterility test methods can
continue to reliably and consistently detect viable contaminating
microorganisms and that verification is the process of confirming that
the sterility test and test components continue to be capable of
consistently detecting viable contaminating microorganisms in the
samples analyzed. In addition, we acknowledge that method suitability
testing using the product is an important part of a validation protocol
for a sterility test method.
3. What information is needed in written procedures for sterility
testing?
    We have finalized, as proposed, the replacement of the requirements
found in current Sec.  610.12(c) entitled Interpretation of test
results, with the requirements that manufacturers must establish,
implement, and follow written procedures for sterility testing. Written
procedures are essential to ensure consistency in sampling, testing,
and interpretation of results and to provide prospective acceptance
criteria for the sterility test. Written procedures should include all
steps to be followed in the sterility test method for initial and
repeat tests and be detailed, clear, and unambiguous. Under the current
good manufacturing practice regulations, manufacturers are required to
document that a drug product satisfactorily conforms to final
specifications for the drug product (Sec.  211.165(a)). As such,
scientifically sound and appropriate specifications, standards,
sampling plans, and test procedures must be designed and written to
ensure that materials conform to appropriate standards of sterility;
and written procedures must include a description of the sampling
method and the number of units per batch to be tested (see Sec.
211.165(c)).
    Under the final rule, manufacturers may use either culture-based or
non-culture-based sterility test methods to evaluate material for
sterility. There are marked differences between culture-based and non-
culture-based sterility tests. Section 610.12(c) provides the minimum
critical considerations that must be included in the written procedures
for culture-based and non-culture-based sterility tests.
    For culture-based sterility test methods, the written procedures
must include, at a minimum, a description of the composition of the
culture media, growth-promotion test requirements, and incubation
conditions (time and temperature). For non-culture-based sterility test
methods, the written procedures must include the composition of test
components, test parameters, including the acceptance criteria, and the
controls used to verify the test method's ability to consistently
detect the presence of viable contaminating microorganisms.
4. What is an appropriate sample for sterility testing?
    Selection of an appropriate sample of a lot is critical for
purposes of sterility testing. Under Sec.  610.12(d) as finalized, due
to the variety of products covered under Sec.  610.12, the regulation
requires that the sample be appropriate to the material being tested.
    (Comment 23) Five comments requested clarification of the proposed
requirement that the sample be ``appropriate to the material being
tested,'' with respect to the size or volume of the final product lot.
The comments asserted that the example provided in the preamble of the
proposed rule, ``For example, a final product lot size of 100,000 units
would necessitate a greater number of samples to be evaluated than a
final product lot size of 5,000 units,'' (76 FR 36019 at 36023),
conflicts with USP Chapter 71 regarding the minimum number of articles
to be tested in relation to the number of articles in the batch.
    (Response) We acknowledge that the example provided in the preamble
of the proposed rule erroneously compared a final product lot size of
100,000 units to one of 5,000 units. We had intended to compare a final
product lot size of 100,000 to one of 500 units. We recognize that this
error may have caused confusion among some readers, and that the
example was inconsistent with the USP Chapter 71 methods for the
minimum number of articles to be tested in relation to the number of
articles in the batch. It was not our intent to suggest that
established USP compendial sterility test methods, including the
minimal number of articles to be tested in relation to the number of
articles in the batch, were unacceptable under the new requirements in
Sec.  610.12(d).
    In order to clarify the new requirement that the sample be
``appropriate to the material being tested,'' we reiterate that in
selecting an appropriate sample size, Sec.  610.12(d) requires that the
following minimal criteria be considered:
     The size or volume of the final product lot. For example,
a final product lot size of 100,000 units would necessitate a greater
number of samples to be evaluated than a final product lot size of 500
units;
     The duration of manufacturing of the drug product.\7\ For
example, it is important that samples be taken at different points of
manufacture, which, at a minimum, should include the beginning, middle,
and end of manufacturing, in an effort to provide evidence of sterility
of the drug product throughout the duration of the manufacturing
process; \8\
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    \7\ See Sec.  210.3(b)(4) for the definition of the term ``drug
product.''
    \8\ See Sec.  211.160(b) for general requirements for laboratory
controls.
---------------------------------------------------------------------------

     The final container configuration and size. We believe
this will ensure appropriate representation of the lot;
     The quantities or concentrations of inhibitors,
neutralizers, and preservatives, if present, in the test material;
     For a culture-based test method, the volume of test
material that results in a dilution of the product that was determined
not to be bacteriostatic or fungistatic; and
     For a non-culture-based test method, the volume of test
material that results in a dilution of the product that does not
inhibit or otherwise hinder the detection of viable contaminating
microorganisms.
    (Comment 24) Two comments stated that the proposed changes related
to sample size are vague and leave too much room for interpretation by
industry as well as investigators or auditors when determining an
appropriate sample size.
    (Response) We disagree that requiring the sample to be appropriate
to the material being tested is vague and leaves too much open to
interpretation. Our intent in requiring that the sample be
``appropriate to the material being tested,'' with consideration of a
list of minimal criteria, is to provide manufacturers flexibility to
retain their existing procedures for sterility testing using culture-
based methods, or to take

advantage of modern methods as they become available, provided that
these modern methods meet certain criteria, as described in our
response to Comment 23. In addition, as noted previously, sterility
test methods are approved by FDA in either a manufacturer's BLA or BLA
supplement, thereby alleviating concern that the final rule leaves too
much room for interpretation.
    (Comment 25) One comment asked FDA to clarify whether the
quantities or concentrations of inhibitors, neutralizers, and
preservatives, if present in the test material, have an impact on
sample size and selection. The comment also asked about the
relationship between the impact of preservatives and any increase in
the sample size.
    (Response) In selecting an appropriate sample size, Sec.  610.12(d)
requires consideration of certain minimal criteria, including the
quantities or concentrations of inhibitors, neutralizers, and
preservatives, if present in the test material. The consideration of
the quantities or concentrations of inhibitors, neutralizers, and
preservatives, if present in the test material, will depend upon the
product and the test method utilized. This provides both manufacturers
of future innovative products, as well as manufacturers of currently
approved products, the flexibility to take advantage of modern methods
or to retain the sterility testing method as approved in the BLA or BLA
supplement.
5. What is required to verify the sterility test?
    As discussed in the preamble to the proposed rule (76 FR 36019 at
36023), verification activities are necessary to demonstrate that
sterility test methods can continue to reliably and consistently detect
viable contaminating microorganisms. The degree of verification that is
necessary depends upon the sterility test method employed. Depending
upon the sterility test method, verification of each individual test
might be appropriate. On the other hand, some sterility test methods
may only need verification activities performed on the selected culture
media or test organisms. Under Sec.  610.12(e), a manufacturer must
perform verification activities appropriate for the sterility test
method chosen, as set forth in the final rule.
    (Comment 26) In the proposed rule (76 FR 36019 at 36020, footnote
6), we proposed to refer to ``growth-promoting properties'' rather than
``growth-promoting qualities'' and requested comments on which term is
most appropriate. We received two comments in response to our request.
Both comments support the use of ``growth-promoting properties'' and
agree that ``growth-promoting properties'' reflects more accurate and
current terminology.
    (Response) We appreciate and agree with these comments and have
retained the term ``growth-promoting properties'' in the final rule.
    (Comment 27) Two comments requested clarification of the
requirements for verification of culture-based test methods. One
comment asked if, for culture-based test methods, all media must
undergo growth-promotion testing over their shelf-life, and if
validation were performed for three lots, whether it is acceptable to
perform growth-promotion testing on the media only when it is initially
received. One comment acknowledged that each media lot would have to be
tested for growth-promotion at least at the beginning and the end of
its use; however, the comment sought clarification whether companies
would be expected to keep performing the test at regular intervals.
    (Response) For culture-based methods, it is important that each lot
of all culture media undergo growth-promotion testing at regular
intervals over the shelf-life of the media, not just when the media is
initially received. The final rule requires that the sterility test and
test components be verified, as appropriate, to demonstrate that they
can continue to consistently detect viable contaminating
microorganisms. The degree of verification depends upon the sterility
test method employed.
    For culture-based test methods, studies must be conducted to
demonstrate that the performance of the test organisms and culture
media are suitable to consistently detect the presence of viable
contaminating microorganisms, including tests for each lot of culture
media to verify its growth-promoting properties over the shelf-life of
the media and not only at the beginning and end of use. Growth-
promotion testing is important to demonstrate that the culture media
are capable of supporting the growth of microorganisms.
    (Comment 28) One comment recommended that with the proposal to
remove the definition of a lot of culture medium currently defined in
Sec.  610.12(e)(2)(i), revisions to the rule should clearly state that
each delivery of each vendor lot of media be ``QC tested'' by the end
user to verify its ability to detect viable microorganisms. The comment
states, ``It must be made clear that the vendor cannot be totally in
control of the product once it has been shipped from the distribution
centre.'' Further, the comment states it is the user's responsibility
to test each delivery of each vendor lot to ensure that undetected
mistreatment of the testing product during its shipment and delivery to
the end-user has not caused deterioration in its efficacy.
    (Response) We agree that the user of the culture media must verify
that each lot can continue to consistently detect viable contaminating
microorganisms. For the reasons noted previously, we do not believe the
suggested changes are needed because the rule, as proposed and now
finalized, already reflects this requirement.
    (Comment 29) One comment stated that usually validation data
provided by the media suppliers are used to cover the shelf-life of the
media and proposed adding the following text ``or media supplier
validation data must be available'' after the text ``over the shelf-
life of the media'' in proposed Sec.  610.12(e)(1) to capture the fact
that the supplier of the media may also supply this parameter.
    (Response) We do not agree that reliance on media supplier
validation data alone, in lieu of testing by the manufacturer, would be
acceptable. Under Sec.  610.12(e)(1) of the final rule, for culture-
based test methods, manufacturers must conduct tests to demonstrate
that the performance of the test organisms and culture media are
suitable to consistently detect the presence of viable contaminating
microorganisms, including tests for each lot of culture media to verify
its growth-promoting properties over the shelf-life of the media.
Therefore, reliance on media supplier validation data alone, in lieu of
testing by the manufacturer, would not be acceptable.
6. Can a sterility test be repeated?
    For the reasons discussed in the preamble to the proposed rule (76
FR 36019 at 36023 to 36024), we have amended the regulations in Sec.
610.12(b) for repeat testing. Therefore, we have eliminated the
reference to repeat testing of bulk material because, under the final
rule, sterility testing is no longer required on bulk material in most
instances. We also have finalized the proposal to eliminate the use of
a second repeat test for final container material to harmonize our
regulatory expectations with current scientific understanding of
quality manufacturing controls.\9\ Under the final rule,

consistent with USP Chapter 71, if the initial test indicates the
presence of microorganisms, then the product being examined does not
comply with the sterility test requirements, unless a thorough
investigation by the quality control unit can conclusively ascribe the
initial evidence of microbial presence to a laboratory error or faulty
materials used in conducting the test.
---------------------------------------------------------------------------

    \9\ See also Barr D., A. Celeste, R. Fish, et al., Application
of Pharmaceutical CGMPs; FDLI (1997) at p. 146 (``In the case of a
clearly identified laboratory error, the retest results substitute
for the original test results. * * * If, on the other hand, no
laboratory error could be identified in the first test, then there
is no scientific basis for discarding the initial out-of-
specification results in favor of passing retest results.'').
---------------------------------------------------------------------------

    If the test of the initial sample is conclusively found to be
invalid, due to laboratory error or faulty test materials, the
sterility test may be repeated one time. If no evidence of
microorganisms is found in the repeat test, the product examined
complies with the test requirements for sterility. If, however,
evidence of microorganisms is found in the repeat test, the product
examined does not comply with the test requirements for sterility.
    Further, as discussed in the preamble to the proposed rule, both a
comparable product that is reflective of the initial sample in terms of
sample location and the stage in the manufacturing process from which
it was taken, and the same sterility test method must be used for both
the initial and repeat tests. This is intended to ensure that the same
volume of material is used for the initial test and each repeat test,
and that the interpretation of the results is conducted in the same
manner.
    (Comment 30) One comment supported FDA's proposal to modify the
provision for repeat testing to harmonize regulatory expectations with
current scientific understanding of quality manufacturing controls by
eliminating the use of a second repeat test of final container material
and agreed with FDA that the proposed modification of the provision for
repeat testing is in accordance with the USP and the European
Pharmacopeia. However, the commenter noted that FDA's proposed
requirement to take repeat test samples that are reflective of the
initial samples may be difficult to fulfill. For instance, the
commenter states, ``* * * at the time when the sterility test might
show a positive result (after a few days), it could be that it is no
longer possible to distinguish which vials were filled at which point
in time.'' The comment suggested deleting the requirement in proposed
Sec.  610.12(f)(3) that the repeat test must be conducted with
``comparable product that is reflective of the initial sample in terms
of sample location and the stage in the manufacturing process from
which it was obtained.''
    (Response) We appreciate the supportive comments. However, we do
not agree with the recommended change to Sec.  610.12(f)(3). We believe
the final rule is consistent with current scientific understanding of
quality manufacturing controls. If a repeat test is conducted, the same
test method must be used for both the initial and repeat tests, and the
repeat test must be conducted with comparable product that is
reflective of the initial sample in terms of sample location and the
stage in the manufacturing process from which it was obtained.
    As discussed in the preamble to the proposed rule, we appreciate
that this final rule could result in the need for some manufacturers to
modify their repeat test procedures. We continue to consider these
modifications to be minor changes in accordance with Sec.  601.12(d)
and to have a minimal potential for an adverse effect on the identity,
strength, quality, purity, or potency of the product as they may relate
to the safety or effectiveness of the product. Therefore, such changes
must be reported in the annual report within 60 days of the anniversary
date of approval of the BLA.
7. What records must be kept relating to sterility testing?
    Previously, Sec.  610.12(h) incorporated by reference the record
keeping and maintenance requirements contained in Sec. Sec.  211.167
and 211.194. We continue to maintain these requirements. As discussed
in the preamble to the proposed rule (76 FR 36019 at 36024), this is
intended to assure that data derived from sterility tests comply with
established specifications. This includes describing the samples
received for testing, stating the method used to test the samples,
identifying the location of relevant validation or verification data,
recording all calculations performed, and stating how the results of
tests performed compare to set specifications.
8. Are there any exceptions to sterility test requirements?
    In the proposed rule we invited comments on whether any of the
current exceptions should be removed (76 FR 36019 at 36024). We
specifically requested comments on whether to remove the exemption for
platelets. Bacterial contamination of platelets is a recognized public
health risk, and the blood collection industry has already called for
and implemented methods to detect and limit or inactivate bacteria in
platelet components. Requiring testing for platelets would be
consistent with these industry practices.
    (Comment 31) In response to our request for comment, a joint
comment from industry groups recommended that FDA continue to except
Whole Blood, Cryoprecipitated Antihemophilic Factor (AHF), Platelets,
Red Blood Cells, and Plasma from the sterility test requirements in
Sec.  610.12. The comment acknowledged that the blood industry has
called for and implemented methods to detect and limit or inactivate
bacteria in platelet components and that some culture-based methods are
in wide use as a quality control tool. However, there are currently no
available tests that will ensure the sterility of platelet products. In
addition, the joint comment noted that if the current exception for
platelets would be removed, manufacturers of blood and blood components
would not be able to satisfy the new requirement. Further, the comment
recommended that FDA vigorously support applications for pathogen
inactivation processes for platelet components. Moreover, the joint
comment noted that any sterility test requirement tied to a BLA is too
narrow an approach to ensure optimal bacterial testing of platelet
products, as any platelet collected or manufactured by a facility that
does not have a BLA would not be subject to the sterility test
regulation. Accordingly, the joint comment recommended that FDA use a
different mechanism to require testing of all platelet products for
bacterial contamination when testing becomes technologically feasible.
    (Response) We appreciate these comments and we generally agree. We
recognize that blood establishments have begun to take steps to test
for bacterial contamination in platelet components. We welcome the
acknowledgement of the importance of bacterial testing and pathogen
inactivation processes for platelet components and believe that
appropriate microbial testing of platelet components may be necessary
to assure product quality. However, while these technologies are
developing, we have retained the exception from this rule for these
products. Instead, we will continue to review these issues and
available technologies and will take appropriate steps at another time
to address microbial testing of blood components.
    (Comment 32) One comment recommended adding an exception stating
that a manufacturer with parametric release programs is not required to
comply with the sterility test requirements. The comment noted that
parametric release for articles sterilized

with moist heat has been recognized by FDA since 1987, and that many
companies have adopted this approach.
    (Response) We disagree with the proposed change and decline to add
an exception for drug products terminally sterilized by moist heat
processes and subject to parametric release because the exception under
Sec.  610.12(h) (previously under Sec.  610.12(g)) already provides for
an exception for such parametric release programs. As noted in FDA's
guidance document entitled ``Guidance for Industry: Submission of
Documentation in Applications for Parametric Release of Human and
Veterinary Drug Products Terminally Sterilized by Moist Heat
Processes,'' dated February 2010, FDA approval of parametric release
must be requested either in an original application submission under 21
CFR 314.50 or 601.2, or in a prior approval supplement under 21 CFR
314.70 or 601.12.
    (Comment 33) Two comments recommended adding other exceptions to
the sterility test requirements. One comment recommended adding
granulocytes to the exception, and one comment recommended adding in
vitro diagnostic devices regulated as biological products, which do not
purport to be sterile.
    (Response) We decline to adopt the suggested changes because
neither granulocytes nor in vitro diagnostic devices, which do not
purport to be sterile, are subject to the sterility test requirements
in Sec.  610.12. Therefore, we believe the recommendations are beyond
the scope of this rule.
    (Comment 34) One comment recommended that the exceptions provision
be revised to ``specifically include or exclude various biological
product types such as Bioequivalent/Biosimilars and combination
products.''
    (Response) We do not believe the suggested change is needed.
Biological products must comply with the applicable requirements in
parts 600 through 680, in addition to other applicable regulations.
    For the reasons discussed in the preamble to the proposed rule (76
FR 36019 at 36024), we have finalized the proposed minor modifications
to the current exception in Sec.  610.12(g)(4)(ii), under which the
Director of CBER or CDER, as appropriate, determines that data
submitted adequately establish that the mode of administration, the
method of preparation, or the special nature of the product precludes
or does not require a sterility test or that the sterility of the lot
is not necessary to assure the safety, purity, and potency of the
product. Specifically, the minor modification that we refer to is the
``route of administration'' rather than the ``mode of administration''
and to ``any other aspect of the product'' rather than ``the special
nature of the product'' in finalized Sec.  610.12(h)(2) so as to
account for novel products that may be introduced to the market in the
future. This exception allows the Director of CBER or CDER, as
appropriate, to exempt biological material from the sterility test
requirements of this section if, based upon the scientific evidence
presented in the BLA or BLA supplement, the data adequately establish
that the route of administration, method of preparation, or any other
aspect of the product precludes or does not necessitate a sterility
test to assure the safety, purity, and potency of the product. We note
that in the proposed rule, the Center for Devices and Radiological
Health was erroneously identified in this exception, instead of CDER.
In the final rule, we have correctly identified CDER in the exception
provision at Sec.  610.12(h)(2).
    In addition to comments regarding exceptions as stated in this
document, we have also eliminated, as proposed, the current exceptions
under Sec.  610.12(g)(1) and (2) because they are no longer necessary
given the flexibility now built into the final rule. In addition, we
have eliminated, as proposed, the current exceptions in Sec.
610.12(g)(5) through (g)(9) because they are no longer necessary and
because the revised rule now requires manufacturers to determine the
appropriate sample volume and size for the material being tested and
requires that the sterility test be ``appropriate to the material being
tested.'' (See 76 FR 36019 at 36024 to 36025 for more information.)

IV. Revisions to Other Regulations

    In addition to the revisions to the sterility regulation in Sec.
610.12, we have also revised, as proposed, two other FDA regulations in
this final rule. These revisions are as follows:
     Section 600.3(q): Previously, Sec.  600.3(q) defined
``sterility'' to mean ``freedom from viable contaminating
microorganisms, as determined by the tests prescribed in Sec.  610.12
of this chapter.'' As proposed, we have reworded this definition to
eliminate the term ``prescribed'' since Sec.  610.12 no longer
prescribes specific test methods. Thus, we have amended Sec.  600.3(q)
to define ``sterility'' as ``freedom from viable contaminating
microorganisms, as determined by tests conducted under Sec.  610.12 of
this chapter.''
     Section 680.3(c) (21 CFR 680.3(c)): As proposed, we have
amended Sec.  680.3(c) to eliminate the term ``prescribed.'' Section
680.3(c) now states that ``A sterility test shall be performed on each
lot of each Allergenic Product, as required by Sec.  610.12 of this
chapter.'' Additionally, we have eliminated Sec.  680.3(c)(1) through
(c)(4) because these exceptions are no longer necessary under the
revisions to Sec.  610.12. (See 76 FR 36019 at 36025 for more
information.)

V. Legal Authority

    FDA is issuing this regulation under the biological products
provisions of the Public Health Service Act (the PHS Act) (42 U.S.C.
262 and 264) and the drugs and general administrative provisions of the
Federal Food, Drug, and Cosmetic Act (the FD&C Act) (sections 201, 301,
501, 502, 503, 505, 510, 701, and 704) (21 U.S.C. 321, 331, 351, 352,
353, 355, 360, 371, and 374). Under these provisions of the PHS Act and
the FD&C Act, we have the authority to issue and enforce regulations
designed to ensure that biological products are safe, effective, pure,
and potent, and to prevent the introduction, transmission, and spread
of communicable disease.

VI. Analysis of Impacts

    FDA has examined the impacts of the final rule under Executive
Order 12866, Executive Order 13563, the Regulatory Flexibility Act (5
U.S.C. 601-612), and the Unfunded Mandates Reform Act of 1996 (Pub. L.
104-4). Executive Orders 12866 and 13563 direct Agencies to assess all
costs and benefits of available regulatory alternatives and, when
regulation is necessary, to select regulatory approaches that maximize
net benefits (including potential economic, environmental, public
health and safety, and other advantages; distributive impacts; and
equity). The Agency believes that this final rule is not a significant
regulatory action under Executive Order 12866.
    The Regulatory Flexibility Act requires Agencies to analyze
regulatory options that would minimize any significant impact of a rule
on small entities. While the rule restricts retesting when sterility
tests are failed, the change codifies an approach for retesting that is
similar to the approach prescribed by the USP. The rule does not
otherwise add any new regulatory responsibilities and generally
increases flexibility for sterility testing. Therefore, the Agency
certifies that the final rule will not have a significant economic
impact on a substantial number of small entities.
    Section 202(a) of the Unfunded Mandates Reform Act of 1995 requires
that Agencies prepare a written statement, which includes an

assessment of anticipated costs and benefits, before proposing ``any
rule that includes any Federal mandate that may result in the
expenditure by State, local, and tribal governments, in the aggregate,
or by the private sector, of $100,000,000 or more (adjusted annually
for inflation) in any one year.'' The current threshold after
adjustment for inflation is $136 million, using the most current (2010)
Implicit Price Deflator for the Gross Domestic Product. FDA does not
expect this final rule to result in any 1-year expenditure that would
meet or exceed this amount.
    These amendments would generally provide manufacturers of
biological products with more flexibility as to how they evaluate the
sterility of their products and reduce the number of evaluations
required. The net effect would be to reduce costs.
    One part of these amendments might impose some additional costs on
manufacturers, however. Under the current regulations, if a biological
product fails a sterility test, the test may be repeated. If the
product passes a subsequent test, it is inferred that the first test
was flawed and only the latter results are used. Under the new
regulations, the test may be repeated only if it is possible to
``ascribe definitively'' the initial failure to ``a laboratory error or
faulty materials used in conducting the sterility testing.''
    This change could increase costs for manufacturers because
additional products could be discarded. The size of the increase, if
any, would be determined by the number of additional lots discarded,
the lot sizes, and the production costs per unit. Some or all of the
costs of this change, could, in turn, be mitigated by the reduction in
losses associated with the provision of contaminated products.
    This change is expected to affect few manufacturers. The method for
sterility testing described in USP Chapter 71 already limits the
repetition of tests to circumstances similar to those described in
these amendments. It is anticipated that, in the absence of these
amendments, the majority of manufacturers would limit the repetition of
sterility tests in order to comply with USP Chapter 71.
    The benefit of limiting retests would be fewer illnesses caused by
contaminated biological products. We are unable to quantify the value
of the reduction in illnesses because we do not have an estimate of the
risk of illness from contaminated biological products or the decline in
that risk associated with limiting retests.

VII. Environmental Impact

    The Agency has determined under 21 CFR 25.31(h) that this action is
of a type that does not individually or cumulatively have a significant
effect on the human environment. Therefore, neither an environmental
assessment nor an environmental impact statement is required.

VIII. Federalism

    FDA has analyzed this final rule in accordance with the principles
set forth in Executive Order 13132. FDA has determined that the rule
does not contain policies that have substantial direct effects on the
States, on the relationship between the National Government and the
States, or on the distribution of power and responsibilities among the
various levels of government. Accordingly, the Agency has concluded
that the rule does not contain policies that have federalism
implications as defined in the Executive order and, consequently, a
federalism summary impact statement is not required.

IX. The Paperwork Reduction Act of 1995

    This final rule contains collections of information that were
submitted for review and approval to the Director of the Office of
Management and Budget (OMB), as required by section 3507(d) of the
Paperwork Reduction Act of 1995 (44 U.S.C. 3501-3520). The collections
of information in Sec. Sec.  211.165 and 610.12 have been approved and
assigned OMB control number 0910-0139.

List of Subjects

21 CFR Part 600

    Biologics, Reporting and recordkeeping requirements.

21 CFR Part 610

    Biologics, Labeling, Reporting and recordkeeping requirements.

21 CFR Part 680

    Biologics, Blood, Reporting and recordkeeping requirements.

    Therefore, under the Federal Food, Drug, and Cosmetic Act, the
Public Health Service Act, and under the authority delegated to the
Commissioner of Food and Drugs, 21 CFR parts 600, 610, and 680 are
amended as follows:

PART 600--BIOLOGICAL PRODUCTS: GENERAL

0
1. The authority citation for 21 CFR part 600 continues to read as
follows:

    Authority: 21 U.S.C. 321, 351, 352, 353, 355, 360, 360i, 371,
374; 42 U.S.C. 216, 262, 263, 263a, 264, 300aa-25.

Sec.  600.3  [Amended]

0
2. Section 600.3 is amended in paragraph (q) by removing ``prescribed
in'' and by adding in its place the phrase ``conducted under''.

PART 610--GENERAL BIOLOGICAL PRODUCTS STANDARDS

0
3. The authority citation for 21 CFR part 610 continues to read as
follows:

    Authority:  21 U.S.C. 321, 331, 351, 352, 353, 355, 360, 360c,
360d, 360h, 360i, 371, 372, 374, 381; 42 U.S.C. 216, 262, 263, 263a,
264.

0
4. Section 610.12 is revised to read as follows:

Sec.  610.12  Sterility.

    (a) The test. Except as provided in paragraph (h) of this section,
manufacturers of biological products must perform sterility testing of
each lot of each biological product's final container material or other
material, as appropriate and as approved in the biologics license
application or supplement for that product.
    (b) Test requirements. (1) The sterility test must be appropriate
to the material being tested such that the material does not interfere
with or otherwise hinder the test.
    (2) The sterility test must be validated to demonstrate that the
test is capable of reliably and consistently detecting the presence of
viable contaminating microorganisms.
    (3) The sterility test and test components must be verified to
demonstrate that the test method can consistently detect the presence
of viable contaminating microorganisms.
    (c) Written procedures. Manufacturers must establish, implement,
and follow written procedures for sterility testing that describe, at a
minimum, the following:
    (1) The sterility test method to be used;
    (i) If culture-based test methods are used, include, at a minimum:
    (A) Composition of the culture media;
    (B) Growth-promotion test requirements; and
    (C) Incubation conditions (time and temperature).
    (ii) If non-culture-based test methods are used, include, at a
minimum:
    (A) Composition of test components;
    (B) Test parameters, including acceptance criteria; and
    (C) Controls used to verify the method's ability to detect the
presence of viable contaminating microorganisms.
    (2) The method of sampling, including the number, volume, and size
of articles to be tested;

    (3) Written specifications for the acceptance or rejection of each
lot; and
    (4) A statement of any other function critical to the particular
sterility test method to ensure consistent and accurate results.
    (d) The sample. The sample must be appropriate to the material
being tested, considering, at a minimum:
    (1) The size and volume of the final product lot;
    (2) The duration of manufacturing of the drug product;
    (3) The final container configuration and size;
    (4) The quantity or concentration of inhibitors, neutralizers, and
preservatives, if present, in the tested material;
    (5) For a culture-based test method, the volume of test material
that results in a dilution of the product that is not bacteriostatic or
fungistatic; and
    (6) For a non-culture-based test method, the volume of test
material that results in a dilution of the product that does not
inhibit or otherwise hinder the detection of viable contaminating
microorganisms.
    (e) Verification. (1) For culture-based test methods, studies must
be conducted to demonstrate that the performance of the test organisms
and culture media are suitable to consistently detect the presence of
viable contaminating microorganisms, including tests for each lot of
culture media to verify its growth-promoting properties over the shelf-
life of the media.
    (2) For non-culture-based test methods, within the test itself,
appropriate controls must be used to demonstrate the ability of the
test method to continue to consistently detect the presence of viable
contaminating microorganisms.
    (f) Repeat test procedures.--(1) If the initial test indicates the
presence of microorganisms, the product does not comply with the
sterility test requirements unless a thorough investigation by the
quality control unit can ascribe definitively the microbial presence to
a laboratory error or faulty materials used in conducting the sterility
testing.
    (2) If the investigation described in paragraph (f)(1) of this
section finds that the initial test indicated the presence of
microorganisms due to laboratory error or the use of faulty materials,
a sterility test may be repeated one time. If no evidence of
microorganisms is found in the repeat test, the product examined
complies with the sterility test requirements. If evidence of
microorganisms is found in the repeat test, the product examined does
not comply with the sterility test requirements.
    (3) If a repeat test is conducted, the same test method must be
used for both the initial and repeat tests, and the repeat test must be
conducted with comparable product that is reflective of the initial
sample in terms of sample location and the stage in the manufacturing
process from which it was obtained.
    (g) Records. The records related to the test requirements of this
section must be prepared and maintained as required by Sec. Sec.
211.167 and 211.194 of this chapter.
    (h) Exceptions. Sterility testing must be performed on final
container material or other appropriate material as defined in the
approved biologics license application or supplement and as described
in this section, except as follows:
    (1) This section does not require sterility testing for Whole
Blood, Cryoprecipitated Antihemophilic Factor, Platelets, Red Blood
Cells, Plasma, Source Plasma, Smallpox Vaccine, Reagent Red Blood
Cells, Anti-Human Globulin, and Blood Grouping Reagents.
    (2) A manufacturer is not required to comply with the sterility
test requirements if the Director of the Center for Biologics
Evaluation and Research or the Director of the Center for Drug
Evaluation and Research, as appropriate, determines that data submitted
in the biologics license application or supplement adequately establish
that the route of administration, the method of preparation, or any
other aspect of the product precludes or does not necessitate a
sterility test to assure the safety, purity, and potency of the
product.

PART 680--ADDITIONAL STANDARDS FOR MISCELLANEOUS PRODUCTS

0
5. The authority citation for 21 CFR part 680 continues to read as
follows:

    Authority:  21 U.S.C. 321, 351, 352, 353, 355, 360, 371; 42
U.S.C. 216, 262, 263, 263a, 264.

0
6. Section 680.3 is amended by revising paragraph (c) to read as
follows:

Sec.  680.3  Tests.

* * * * *
    (c) Sterility. A sterility test shall be performed on each lot of
each Allergenic Product as required by Sec.  601.12 of this chapter.

    Dated: April 27, 2012.
Leslie Kux,
Assistant Commissioner for Policy.
[FR Doc. 2012-10649 Filed 5-2-12; 8:45 am]
BILLING CODE 4160-01-P