Analytical Procedures 
and Methods Validation 
for Drugs and Biologics 
 
 
Guidance for Industry 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Drug Evaluation and Research (CDER) 
Center for Biologics Evaluation and Research (CBER) 
 
July 2015 
Pharmaceutical Quality/CMC 
 
   
Analytical Procedures 
and Methods Validation 
for Drugs and Biologics 
Guidance for Industry 
 
Additional copies are available from: 
Office of Communications, Division of Drug Information  
Center for Drug Evaluation and Research 
Food and Drug Administration 
10001 New Hampshire Ave., Hillandale Bldg., 4th Floor  
Silver Spring, MD 20993  
Phone: 855-543-3784 or 301-796-3400; Fax: 301-431-6353 
Email: druginfo@fda.hhs.gov  
http://www.fda.gov/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/default.htm 
and/or 
Office of Communication, Outreach and Development  
Center for Biologics Evaluation and Research 
 Food and Drug Administration 
10903 New Hampshire Ave., Bldg. 71, Room 3128 
Silver Spring, MD 20993 
Phone: 800-835-4709 or 240-402-7800 
Email: ocod@fda.hhs.gov 
http://www.fda.gov/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInformation/Guidances/default.htm    
 
 
 
 
 
 
 
 
 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Drug Evaluation and Research (CDER) 
Center for Biologics Evaluation and Research (CBER) 
 
July 2015 
Pharmaceutical Quality/CMC 
 
  Contains Nonbinding Recommendations 
TABLE OF CONTENTS 
 
 
I.  INTRODUCTION............................................................................................................. 1 
II.  BACKGROUND ............................................................................................................... 2 
III.  ANALYTICAL METHODS DEVELOPMENT ............................................................ 3 
IV.  CONTENT OF ANALYTICAL PROCEDURES .......................................................... 4 
A.  Principle/Scope ............................................................................................................................... 4 
B.  Apparatus/Equipment ................................................................................................................... 4 
C.  Operating Parameters ................................................................................................................... 4 
D.  Reagents/Standards ....................................................................................................................... 4 
E.  Sample Preparation ....................................................................................................................... 5 
F.  Standards Control Solution Preparation ..................................................................................... 5 
G.  Procedure ........................................................................................................................................ 5 
H.  System Suitability .......................................................................................................................... 5 
I.  Calculations .................................................................................................................................... 5 
J.  Data Reporting ............................................................................................................................... 6 
V.  REFERENCE STANDARDS AND MATERIALS........................................................ 6 
VI.  ANALYTICAL METHOD VALIDATION ................................................................... 7 
A.  Noncompendial Analytical Procedures ........................................................................................ 7 
B.  Validation Characteristics............................................................................................................. 7 
C.  Compendial Analytical Procedures .............................................................................................. 8 
VII.  STATISTICAL ANALYSIS AND MODELS ................................................................ 8 
A.  Statistics .......................................................................................................................................... 8 
B.  Models ............................................................................................................................................. 9 
VIII. LIFE CYCLE MANAGEMENT OF ANALYTICAL PROCEDURES ........................ 9 
A.  Revalidation .................................................................................................................................. 10 
B.  Analytical Method Comparability Studies ................................................................................ 10 
1. Alternative Analytical Procedures ................................................................................................. 10 
2. Analytical Methods Transfer Studies ............................................................................................. 12 
C.  Reporting Postmarketing Changes to an Approved NDA, ANDA, or BLA ........................... 12 
IX.  FDA METHODS VERIFICATION .............................................................................. 12 
X.   REFERENCES ................................................................................................................ 13 
  Contains Nonbinding Recommendations 
Analytical Procedures and Methods Validation for Drugs and 
1 
Biologics 
2 
1
Guidance for Industry  
3 
4   
5   
6  This guidance represents the current thinking of the Food and Drug Administration (FDA or Agency) on 
7  this topic.  It does not create any rights for any person and is not binding on FDA or the public.  You can 
8  use an alternative approach if it satisfies the requirements of the applicable statutes and regulations.  To 
9  discuss an alternative approach, contact the FDA staff responsible for this guidance as listed on the title 
10  page.   
11   
12   
13   
14  I.  INTRODUCTION  
15   
16  This guidance supersedes the draft of the same name that published on February 19, 2014 (79 FR 
17  9467)  and replaces the 2000 draft guidance for industry on Analytical Procedures and Methods 
18  Validation2,3 and the 1987 Guidelines for Submitting Samples and Analytical Data for Methods 
19  Validation. It provides recommendations on how you, the applicant, can submit analytical 
20  procedures4 and methods validation5 data to support the documentation of the identity, strength, 
21  quality, purity, and potency of drug substances and drug products.6  It will help you assemble 
22  information and present data to support your analytical methodologies.  The recommendations 
23  apply to drug substances and drug products covered in new drug applications (NDAs), 
24  abbreviated new drug applications (ANDAs), biologics license applications (BLAs), and 
25  supplements to these applications.  The principles in this guidance also apply to drug substances 
26  and drug products covered in Type II drug master files (DMFs).  
27   
28  This guidance complements the International Conference on Harmonisation (ICH) guidance 
29  Q2(R1) Validation of Analytical Procedures:  Text and Methodology (Q2(R1)) for developing and 
30  validating analytical methods.    
31   
32  This guidance does not address investigational new drug application (IND) methods validation, 
33  but sponsors preparing INDs should consider the recommendations in this guidance.  For INDs, 
34  sufficient information is required at each phase of an investigation to ensure proper identity, 
35  quality, purity, strength, and/or potency.  The amount of information on analytical procedures 
36  and methods suitability will vary with the phase of the investigation.7  For general guidance on 
                                                           
1 This guidance has been prepared by the Office of Pharmaceutical Quality in the Center for Drug Evaluation and 
Research (CDER) and the Center for Biologics Evaluation and Research (CBER) at the Food and Drug 
Administration.  
2 Sample submission is described in section IX, FDA Methods Verification.    
3 We update guidances periodically.  To make sure you have the most recent version of a guidance, check the FDA 
Drugs guidance Web page at 
http://www.fda.gov/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/default.htm.  
4 Analytical procedure is interchangeable with a method or test procedure. 
5 Compendial methods are verified rather than validated as described in section VI, C. 
6 The terms drug substance and drug product are used in this guidance to refer to both human drugs and biologics. 
7  See 21 CFR 312.23(a)(7). 
  1  Contains Nonbinding Recommendations 
37  analytical procedures and methods validation information to be submitted for phase one studies, 
38  sponsors should refer to the FDA guidance for industry on Content and Format of 
39  Investigational New Drug Applications (INDs) for Phase 1 Studies of Drugs, Including 
40  Well-Characterized, Therapeutic, Biotechnology-Derived Products.  General considerations for 
41  analytical procedures and methods validation before conduct of phase two and three studies are 
42  discussed in the FDA guidances for industry on INDs for Phase 2 and 3 Studies of Drugs, 
43  Including Specified Therapeutic Biotechnology-Derived Products (February 1999) and IND 
44  Meetings for Human Drugs and Biologics, Chemistry, Manufacturing, and Controls 
45  Information. 
46   
47  This guidance does not address specific method validation recommendations for biological and 
48  immunochemical assays for characterization and quality control of many drug substances and 
49  drug products.  For example, some bioassays are based on animal challenge models, and 
50  immunogenicity assessments or other immunoassays have unique features that should be 
51  considered during development and validation.  
52   
53  Analytical methods required during product and process development activities are discussed in FDA 
54  guidance for industry on Process Validation: General Principles and Practices. 
55   
56  In addition, a risk-based approach on the need for revalidation of existing analytical methods 
57  may need to be considered when the manufacturing process changes during the products life 
58  cycle.  For questions on appropriate validation approaches for analytical procedures or 
59  submission of information not addressed in this guidance, you should consult with the 
60  appropriate FDA quality assessment staff. 
61   
62  If you choose a different approach than those recommended in this guidance, we encourage you 
63  to discuss the matter with the appropriate FDA quality assessment staff before you submit your 
64  application.  
65   
66  In general, FDAs guidance documents do not establish legally enforceable responsibilities.  
67  Instead, guidances describe the Agencys current thinking on a topic and should be viewed only 
68  as recommendations, unless specific regulatory or statutory requirements are cited.  The use of 
69  the word should in Agency guidances means that something is suggested or recommended, but 
70  not required.  
71   
72   
73  II.  BACKGROUND 
74   
75  Each NDA and ANDA must include the analytical procedures necessary to ensure the identity, 
76  strength, quality, purity, and potency of the drug substance and drug product.8  Each BLA must 
77  include a full description of the manufacturing process, including analytical procedures that 
78  demonstrate the manufactured product meets prescribed standards of identity, quality, safety, 
79  purity, and potency.9  Data must be available to establish that the analytical procedures used in 
                                                           
8  See 21 CFR 314.50(d)(1) and 314.94(a)(9)(i). 
9  See 21 CFR 601.2(a) and 601.2(c). 
  2  Contains Nonbinding Recommendations 
80  testing meet proper standards of accuracy, sensitivity, specificity, and reproducibility and are 
81  suitable for their intended purpose.10   
82   
83  Analytical procedures verification or validation data should be submitted in the corresponding 
84  sections of the application in the ICH M2 eCTD: Electronic Common Technical Document 
85  Specification.11 
86   
87  When an analytical procedure is approved/licensed as part of the NDA, ANDA, or BLA, it 
88  becomes the FDA-approved analytical procedure for the approved product.  This analytical 
89  procedure may originate from FDA recognized sources (e.g., a compendial procedure from the 
90  United States Pharmacopeia/National Formulary (USP/NF)) or a validated procedure you 
91  submitted that was determined to be acceptable by FDA.  To apply an analytical method to a 
92  different drug product, appropriate validation or verification studies for compendial procedures 
93  with the matrix of the new product should be considered. 
94   
95   
96  III.  ANALYTICAL METHODS DEVELOPMENT 
97   
98  An analytical procedure is developed to test a defined characteristic of the drug substance or 
99  drug product against established acceptance criteria for that characteristic.  Early in the 
100  development of a new analytical procedure, the choice of analytical instrumentation and 
101  methodology should be selected based on the intended purpose and scope of the analytical 
102  method.  Parameters that may be evaluated during method development are specificity, linearity, 
103  limits of detection (LOD) and limits of quantitation (LOQ), range, accuracy, and precision.  
104   
105  During early stages of method development, the robustness of methods should be evaluated 
106  because this characteristic can help you decide which method you will submit for approval.  
107  Analytical procedures in the early stages of development are initially developed based on a 
108  combination of mechanistic understanding of the basic methodology and prior experience.  
109  Experimental data from early procedures can be used to guide further development.  You should 
110  submit development data within the method validation section if they support the validation of 
111  the method. 
112   
113  To fully understand the effect of changes in method parameters on an analytical procedure, you 
114  should adopt a systematic approach for a method robustness study (e.g., a design of experiments 
115  with method parameters).  You should begin with an initial risk assessment and follow with 
116  multivariate experiments.  Such approaches allow you to understand factorial parameter effects 
117  on method performance.  Evaluation of a methods performance may include analyses of 
118  samples obtained from various stages of the manufacturing process from in-process to the 
119  finished product.  Knowledge gained during these studies on the sources of method variation can 
120  help you assess the method performance.   
121   
122   
                                                           
10 See 21 CFR 211.165(e) and 211.194(a)(2). 
11 Sections as applicable in Module 3: 3.2.S and 3.2.P. 
  3  Contains Nonbinding Recommendations 
123  IV.  CONTENT OF ANALYTICAL PROCEDURES 
124   
125  You should describe analytical procedures in sufficient detail to allow a competent analyst to 
126  reproduce the necessary conditions and obtain results within the proposed acceptance criteria.  
127  You should also describe aspects of the analytical procedures that require special attention.  An 
128  analytical procedure may be referenced from FDA-recognized sources (e.g., USP/NF, 
129  Association of Analytical Communities (AOAC) International)12 if the referenced analytical 
130  procedure is not modified beyond what is allowed in the published method.  You should provide 
131  in detail procedures from other published sources.  The following is a list of essential 
132  information you should include for an analytical procedure:  
133   
134  A.  Principle/Scope 
135   
136  A description of the basic principles of the analytical test/technology (i.e., separation, detection); 
137  target analyte(s) and sample(s) type (e.g., drug substance, drug product, impurities or compounds 
138  in biological fluids).   
139   
140  B.  Apparatus/Equipment 
141   
142  All required qualified equipment and components  e.g., instrument type, detector, column type, 
(
143  dimensions, and alternative column, filter type). 
144   
145  C.  Operating Parameters 
146   
147  Qualified optimal settings and ranges (include allowed adjustments supported by compendial 
148  sources or development and/or validation studies) critical to the analysis (e.g., flow rate, 
149  components temperatures, run time, detector settings, gradient, head space sampler).  A drawing 
150  with experimental configuration and integration parameters may be used, as applicable.    
151   
152  D.  Reagents/Standards 
153   
154  The following should be listed where applicable: 
155   
156    Description of reagent or standard  
157    Grade of chemical (e.g., USP/NF, American Chemical Society, High 
158  Performance or Pressure Liquid Chromatography, or Gas 
159  Chromatography and preservative-free)  
160    Source (e.g., USP reference standard, qualified in-house reference material, 
161  WHO International Standard/Reference Material, CBER standard)  
162    Purity (for pure chemicals only), State (e.g., dried, undried), and concentration  
163    Potencies (where required by CFR, USP)  
164    Storage conditions  
165    Directions for safe use (as per current Safety Data Sheet)  
166    Validated or documented shelf life   
                                                           
12  See 21 CFR 211.194(a)(2). 
  4  Contains Nonbinding Recommendations 
167   
168  New batches of biological reagents, such as monoclonal antibodies, polyclonal antisera, or cells, 
169  may need extensive qualification procedures included as part of the analytical procedure. 
170   
171  E.  Sample Preparation 
172   
173  Procedures (e.g., extraction method, dilution or concentration, desalting procedures and mixing 
174  by sonication, shaking or sonication time) for the preparations for individual sample tests.  A 
175  single preparation for qualitative and replicate preparations for quantitative tests with appropriate 
176  units of concentrations for working solutions (e.g., g/ml or mg/ml) and information on stability 
177  of solutions and storage conditions. 
178   
179  F.  Standards Control Solution Preparation 
180   
181  Procedures for the preparation and use of all standard and control solutions with appropriate 
182  units of concentration and information on stability of standards and storage conditions,   
183  including calibration standards, internal standards, system suitability standards, etc.  
184   
185  G.  Procedure 
186   
187  A step-by-step description of the method (e.g., equilibration times, and scan/injection sequence 
188  with blanks, placeboes, samples, controls, sensitivity solution (for impurity method) and 
189  standards to maintain validity of the system suitability during the span of analysis) and allowable 
190  operating ranges and adjustments if applicable.  
191   
192  H.  System Suitability 
193   
194  Confirmatory test(s) procedures and parameters to ensure that the system (equipment, 
195  electronics, and analytical operations and controls to be analyzed) will function correctly as an 
196  integrated system at the time of use.  The system suitability acceptance criteria applied to 
197  standards controls and samples, such as peak tailing, precision and resolution acceptance criteria, 
198  may be required as applicable.  For system suitability of chromatographic systems, refer to the 
199  FDA guidance for industry on Validation of Chromatographic Methods and USP General 
200  Chapter <621> Chromatography.  
201    
202  I.  Calculations 
203   
204  The integration method and representative calculation formulas for data analysis (standards, 
205  controls, samples) for tests based on label claim and specification (e.g., assay, specified and 
206  unspecified impurities and relative response factors).  This includes a description of any 
207  mathematical transformations or formulas used in data analysis, along with a scientific 
208  justification for any correction factors used. 
209   
  5  Contains Nonbinding Recommendations 
210  J.  Data Reporting  
211   
212  A presentation of numeric data that is consistent with instrumental capabilities and acceptance 
213  criteria.  The method should indicate what format to use to report results (e.g., percentage label 
214  claim, weight/weight, and weight/volume) with the specific number of significant figures 
215  needed.  The American Society for Testing and Materials (ASTM) E29 standard describes a 
216  standard practice for using significant digits in test data to determine conformance with 
217  specifications. For chromatographic methods, you should include retention times (RTs) for 
218  identification with reference standard comparison basis, relative retention times (RRTs) (known 
219  and unknown impurities) acceptable ranges and sample results reporting criteria. 
220   
221   
222  V.  REFERENCE STANDARDS AND MATERIALS 
223   
224  Primary and secondary reference standards and materials are defined and discussed in the 
225  following ICH guidances:  Q6B Specifications:  Test Procedures and Acceptance Criteria for 
226  Biotechnological/Biological Products, and Q7 Good Manufacturing Practice Guidance for 
227  Active Pharmaceutical Ingredients.  For all standards, you should ensure the suitability for use.  
228  You should strictly follow storage and usage conditions and handling instructions for reference 
229  standards to avoid modifications and contaminations, which could result in additional impurities 
230  and inaccurate analysis.  You should include information supporting any reference standards and 
231  materials that you intend to use in the application. Information supporting reference standards 
232  and materials should include qualification test reports and certificates of analysis (including 
233  stability protocols, reports, and relevant known impurity profile information) as applicable. For 
234  biological products under BLAs, qualification of subsequent reference standard lots should be 
235  included in annual reports. 
236   
237  Reference standards can often be obtained from USP and may also be available through the 
238  European Pharmacopoeia, Japanese Pharmacopoeia, World Health Organization, or National 
239  Institute of Standards and Technology.  Reference standards for a number of biological products 
240  are also available from CBER.  For certain biological products marketed in the U.S., reference 
241  standards authorized by CBER must be used before the product can be released to the market.13  
242  Reference materials from other sources should be characterized by procedures including routine 
243  and beyond routine release testing as described in ICH Q6B.  You should consider orthogonal 
244  methods for reference material characterization.  Additional testing could include attributes to 
245  determine the suitability of the reference material not necessarily captured by the drug substance 
246  or product release tests (e.g., more extensive structural identity and orthogonal techniques for 
247  potency, purity and impurities). 
248   
249  A new batch of reference standard material (official or in-house) should be qualified/calibrated 
250  against the current reference standard.  For biological reference standards and materials, we 
251  recommend that you follow a two-tiered approach when qualifying new reference standards to 
252  prevent drift in the quality attributes. A two-tiered approach involves a comparison of each new 
                                                           
13 See 21 CFR 610.20. 
 
  6  Contains Nonbinding Recommendations 
253  reference standard with a primary reference standard so that it is linked to clinical trial material 
254  and the current manufacturing process. 
255   
256   
257  VI.  ANALYTICAL METHOD VALIDATION  
258   
259  A.  Noncompendial Analytical Procedures 
260   
261  Analytical method validation is the process of demonstrating that an analytical procedure is 
262  suitable for its intended purpose.  The methodology and objective of the analytical procedures 
263  should be clearly defined and understood before initiating validation studies.  This understanding 
264  is obtained from scientifically-based method development and optimization studies.  Validation 
265  data must be generated under a protocol approved by the sponsor following current good 
266  manufacturing practices with the description of methodology of each validation characteristic 
267  and predetermined and justified acceptance criteria, using qualified instrumentation.14  Protocols 
268  for both drug substance and product analytes or mixture of analytes in respective matrices should 
269  be developed and executed.  You should include details of the validation studies and results with 
270  your application.  
271   
272  B.  Validation Characteristics 
273   
274  Although not all of the validation characteristics are applicable for all types of tests, typical 
275  validation characteristics are: 
276   
277    Specificity  
278    Linearity 
279    Accuracy  
280    Precision (repeatability, intermediate precision, and reproducibility) 
281    Range 
282    Quantitation limit 
283    Detection limit 
284   
285  ICH Q2(R1) is considered the primary reference for recommendations and definitions on 
286  validation characteristics for analytical procedures.  The FDA guidance for industry on  
287  Validation of Chromatographic Methods is available as well. 
288   
289  If a procedure is a validated quantitative analytical procedure that can detect changes in a quality 
290  attribute(s) of the drug substance and drug product during storage, it is considered a stability-
291  indicating test.  To demonstrate specificity of a stability-indicating test, a combination of 
292  challenges should be performed.  Some challenges include the use of samples spiked with target 
293  analytes and all known interferences; samples that have undergone various laboratory stress 
294  conditions; and actual product samples (produced by the final manufacturing process) that are 
295  either aged or have been stored under accelerated temperature and humidity conditions. 
296   
                                                           
14 For drugs see 21 CFR 211.165(e); 21 CFR 314.50 (d), and for biologics see 21 CFR 601.2(a), 601.2(c), and 
601.12(a). 
  7  Contains Nonbinding Recommendations 
297  As the holder of the NDA, ANDA, or BLA, you must: (1) submit the data used to establish that 
298  the analytical procedures used in testing meet proper standards of accuracy and reliability, and 
299  (2) notify the FDA about each change in each condition established in an approved application 
300  beyond the variations already provided for in the application, including changes to analytical 
301  procedures and other established controls.15 
302   
303  The submitted data should include the results from the robustness evaluation of the method, 
304  which is typically conducted during method development or as part of a planned validation 
305  study.16 
306   
307  C.  Compendial Analytical Procedures 
308   
309  The suitability of an analytical procedure (e.g., USP/NF, the Official Methods of Analysis of 
310  AOAC International, or other recognized standard references) should be verified under actual 
311  conditions of use.17  Information to demonstrate that USP/NF analytical procedures are suitable 
312  for the drug product or drug substance should be included in the submission and generated under 
313  a verification protocol.   
314   
315  The verification protocol should include, but is not limited to:  (1) compendial methodology to 
316  be verified with predetermined acceptance criteria, and (2) details of the methodology (e.g., 
317  suitability of reagent(s), equipment, component(s), chromatographic conditions, column, detector 
318  type(s), sensitivity of detector signal response, system suitability, sample preparation and 
319  stability).  The procedure and extent of verification should dictate which validation characteristic 
320  tests should be included in the protocol (e.g., specificity, LOD, LOQ, precision, accuracy). 
321  Considerations that may influence what characteristic tests should be in the protocol may depend 
322  on situations such as whether specification limits are set tighter than compendial acceptance 
323  criteria, or RT or RRT profiles are changing in chromatographic methods because of the 
324  synthetic route of drug substance or differences in manufacturing process or matrix of drug 
325  product.  Robustness studies of compendial assays do not need to be included, if methods are 
326  followed without deviations. 
327   
328   
329  VII.  STATISTICAL ANALYSIS AND MODELS 
330   
331  A.  Statistics  
332   
333  Statistical analysis of validation data can be used to evaluate validation characteristics against 
334  predetermined acceptance criteria.  All statistical procedures and parameters used in the analysis 
335  of the data should be based on sound principles and appropriate for the intended evaluation.  
336  Several statistical methods are useful for assessing validation characteristics, for example, an 
337  analysis of variance (ANOVA) to assess regression analysis R (correlation coefficient) and R 
                                                           
15 For drugs see 21 CFR 314.50 (d), 314.70(d), and for biologics see 21 CFR 601.2(a), 601.2(c), and 601.12(a). For a 
BLA, as discussed, you must obtain prior approval from FDA before implementing a change in analytical methods if 
those methods are specified in FDA regulations. 
16 See section III and ICH Q2(R1). 
17 See 21 CFR 211.194(a)(2) and USP General Chapter <1226> Verification of Compendial Procedures. 
  8  Contains Nonbinding Recommendations 
338  squared (coefficient of determination) or linear regression to measure linearity.  Many statistical 
339  methods used for assessing validation characteristics rely on population normality, and it is 
340  important to determine whether or not to reject this assumption.  There are many techniques, 
341  such as histograms, normality tests, and probability plots that can be used to evaluate the 
342  observed distribution.  It may be appropriate to transform the data to better fit the normal 
343  distribution or apply distribution-free (nonparametric) approaches when the observed data are 
344  not normally distributed.  Appropriate literature or text should be consulted for information on 
345  statistical procedures to use when developing new test methods, evaluating existing test methods 
346  or evaluating measurement system performance, as well as other general information on the 
347  interpretation and treatment of analytical data.18  The data analysis should be assured either by 
348  using appropriately validated software or independent verification for correctness.  
349    
350  B.  Models 
351   
352  Some analytical methods might use chemometric and/or multivariate models.  When developing 
353  these models, the number of samples to provide adequate statistical power and range for model 
354  development and validation should be considered.  Suitable software should be used for data 
355  analysis.  Model parameters should be deliberately varied to test model robustness.  
356   
357   
358  VIII. LIFE CYCLE MANAGEMENT OF ANALYTICAL PROCEDURES 
359   
360  Once an analytical procedure (including compendial methods) is successfully validated (or 
361  verified) and implemented, the procedure should be followed during the life cycle of the product 
362  to continually assure that it remains fit for its intended purpose.  Trend analysis on method 
363  performance should be performed at regular intervals to evaluate the need to optimize the 
364  analytical procedure or to revalidate all or a part of the analytical procedure.  If an analytical 
365  procedure can only meet the established system suitability requirements with repeated 
366  adjustments to the operating conditions stated in the analytical procedure, the analytical 
367  procedure should be reevaluated, revalidated, or amended, as appropriate.   
368   
369  Over the life cycle of a product, new information and risk assessments (e.g., a better 
370  understanding of product CQAs or awareness of a new impurity) may warrant the development 
371  and validation of a new or alternative analytical method.  New technologies may allow for 
372  greater understanding and/or confidence when ensuring product quality.  Applicants should 
373  periodically evaluate the appropriateness of a products analytical methods and consider new or 
374  alternative methods. 
375   
376  In anticipation of life cycle changes in analytics, an appropriate number of retention samples 
377  should be maintained to allow for comparative studies.  The number should be based on 
378  scientific principles and an assessment of risk. For complex products that are sensitive to 
379  manufacturing changes, reserve samples can be an important tool to make these comparisons.  
                                                           
18 See References section for examples including USP <1010> Analytical Data  Interpretation and Treatment, 
ASTM E1488 Standard Guide for Statistical Procedures to Use in Developing and Applying Test Methods and 
ASTM E2782 Standard Guide for Measurement Systems Analysis. 
  9  Contains Nonbinding Recommendations 
380  The retention samples used in comparative studies should include samples that represent 
381  marketed product and, when possible, pivotal clinical trial material.  
382     
383  If a risk-based evaluation or other drivers lead to changes in an analytical procedure or 
384  replacement with a new method or if the procedure is transferred to a new testing site; 
385  revalidation, a new validation exercise, an analytical method comparability study, or a 
386  combination of these exercises should be considered.  In some cases, changes to the drug 
387  substance or drug product manufacturing process may also warrant analytical procedure 
388  revalidation.  These additional studies are discussed below. 
389   
390  A.  Revalidation  
391   
392  Principles described in the validation section (section VI) apply to revalidation. When a change 
393  is made to an analytical procedure (e.g., a change in a piece of equipment or reagent or because 
394  of a change in manufacturing process or formulation), revalidation of all or part of the analytical 
395  procedure should be considered.  Analytical method revalidation may also be warranted because 
396  of manufacturing process changes, such as an alteration in the drug substance manufacturing 
397  process that could impact method performance (e.g., route of synthesis, fermentation) or 
398  introduction of a new drug product formulation. 
399   
400  You should revalidate to ensure that the analytical procedure maintains its critical performance 
401  characteristics (e.g., specificity, precision, accuracy).  The degree of revalidation depends on the 
402  nature of the change.  
403   
404  B.  Analytical Method Comparability Studies 
405   
406  Analytical method comparability study requests are typically generated when you propose to 
407  substitute an FDA-approved analytical procedure with an alternative analytical procedure or 
408  when an analytical method is transferred from one laboratory to the other.  For information on 
409  statistical procedures to use for determining equivalence of two test methods, appropriate 
410  literature or text should be consulted.19  These scenarios are discussed below. 
411   
412  1.  Alternative Analytical Procedures 
413   
414  An alternative analytical procedure is an analytical procedure that you use in place of the FDA-
415  approved analytical procedure.  For an NDA or ANDA, you should include any proposed 
416  alternate analytical procedures in the application. You must include a description of the 
417  procedure.20  After approval, for an NDA or ANDA, or for a procedure approved in a BLA but 
418  not included in an FDA regulation, the addition, revision, or deletion of an alternative analytical 
419  procedure that provides the same or increased assurance of the identity, strength, quality, purity, 
                                                           
19 See References section for examples including USP General Chapter <1010> Analytical Data  Interpretation 
and Treatment and ASTM E2935 Standard Practice for Conducting Equivalence Testing in Laboratory 
Applications. 
20 See 21 CFR 314.50. 
  10  Contains Nonbinding Recommendations 
420  or potency of the material being tested as the analytical procedure described in the approved 
421  application, must be documented in the next annual report.21  
422   
423  For biological products, in rare cases an analytical procedure may be included in an FDA 
424  regulation.  If the analytical method required is described by a regulation, however, and you want 
425  to use an alternate method, you must submit the alternate method for review and approval 
426  according to 21 CFR 610.9(a).  You must present evidence ...demonstrating that the 
427  modification will provide assurances of the safety, purity, potency, and effectiveness of the 
428  biological product equal to or greater than the assurances provided by the method or process 
429  specified in the general standards or additional standards for the biological product.  
430  Modification of such procedures requires FDA approval during application review or in a 
431  postapproval supplement.22 
432   
433  You should identify the use of the alternative analytical procedure (e.g., release, stability testing) 
434  and provide a rationale for its inclusion, validation data, and comparative data to the FDA-
435  approved analytical procedure.  You should perform an analytical method comparability study 
436  that demonstrates at a minimum that:   
437   
438    The new method coupled with any additional control measures is equivalent or 
439  superior to the original method for the intended purpose.  
440   
441    The new analytical procedure is not more susceptible to matrix effects than the 
442  original procedure. 
443   
444  If new process-related or product-related variants or any new impurities are discovered with the 
445  new procedure, testing on retention samples from historical batches should be performed to 
446  demonstrate that the variants/impurities detected by the new method are a result of an increase in 
447  the sensitivity or selectivity of the new procedure and not a result of a change to process-related 
448  impurities.  
449   
450  If the procedure has stability-indicating properties: 
451   
452    Appropriate samples should be included that allow a comparison of the ability of 
453  the new and original method to detect relevant product variants and degradation 
454  species.   
455    The number of batches analyzed for comparison should provide sufficient 
456  statistical power.    
457    Equivalence, non-inferiority, or superiority studies should be performed with 
458  appropriate statistical methods to demonstrate that the new or revised methods 
459  performance is comparable or better than the original method. 23   
460    The statistical analyses performed to compare product testing should be 
461  identified.   
                                                           
21 See 21 CFR 314.70(d)(1), (d)(2)(vii).  314.81(b)(2), and 601.12(d)(vii).   
22 See 21 CFR 610.9(b). 
23 ASTM E2935  Standard Practice for Conducting Equivalence Testing in Laboratory Applications. 
  11  Contains Nonbinding Recommendations 
462    All bias or differences between analytical procedures seen with comparative 
463  results should be discussed with an explanation, as appropriate. 
464   
465  2.  Analytical Methods Transfer Studies 
466   
467  Analytical method transfer is typically managed under a transfer protocol that details the 
468  parameters to be evaluated in addition to the predetermined acceptance criteria that will be 
469  applied to the results.  Transfer studies usually involve two or more laboratories or sites 
470  (originating lab and receiving labs) executing the preapproved transfer protocol.  A sufficient 
471  number of representative test articles (e.g., same lot(s) of drug substance or drug product) are 
472  used by the originating and receiving laboratories.  The comparative studies are performed to 
473  evaluate accuracy and precision, especially with regard to assessment of interlaboratory 
474  variability.  In cases where the transferred analytical procedure is also a stability-indicating 
475  method, forced degradation samples or samples containing pertinent product-related impurities 
476  should be analyzed at both sites.  The USP General Chapter <1224> Transfer of Analytical 
477  Procedures provides additional guidance on this topic. 
478   
479  C.  Reporting Postmarketing Changes to an Approved NDA, ANDA, or BLA 
480   
481  Postmarketing changes to analytical procedures must be reported to the FDA in compliance with 
482  21 CFR 314.70 or 21 CFR 601.12.24  Additional information on the appropriate reporting 
483  category for various kinds of postapproval changes for NDAs and ANDAs is provided in the 
484  FDA guidance for industry on Changes to an Approved NDA or ANDA and Changes to an 
485  Approved NDA or ANDA; Specifications  Use of Enforcement Discretion for Compendial 
486  Changes.  Similar information on postapproval changes to BLAs regulated by CDER and CBER 
487  is provided in the FDA guidance Changes to an Approved Application for Specified 
488  Biotechnology and Specified Synthetic Biological Products. 
489   
490   
491  IX.  FDA METHODS VERIFICATION  
492   
493  Part of the approval process for NDAs and ANDAs may include FDA laboratory assessment to 
494  determine whether the analytical procedures are acceptable for quality control and suitable for 
495  regulatory purposes.25  If a laboratory assessment will be conducted, the FDA laboratory will 
496  send you a request that will detail what samples and supplies to send to the FDA laboratory.  
497  These could include product samples, standards, critical reagents, material safety data sheets, and 
498  supplies.  Laboratory results and comments will be forwarded from the FDA laboratory to the 
499  product quality reviewer.  
500   
501  For certain biological products, samples representative of the product for licensure along with 
502  summaries of results of tests performed on the lots represented by these samples should be 
503  submitted with the BLA.26  The FDA laboratory verifies the performance of the methods and the 
                                                           
24 As noted, for a product licensed under a BLA, if the change is to a procedure prescribed in FDA regulations that 
change must be approved by FDA pursuant to 21 CFR 610.9(b). 
25 See 21 CFR 314.50(e). 
26 See 21 CFR 601.2(a). 
  12  Contains Nonbinding Recommendations 
504  results you submit. During a pre-BLA meeting or after submission of the BLA, the FDA 
505  laboratory can send you a request to provide standards, controls, reagents, material safety data 
506  sheets, and supplies.   
507   
508  X.   REFERENCES 
509     
510  Guidance for Industry27 
511   
512  ANDAs:  Impurities in Drug Products (November 2010) 
513   
514  ANDAs:  Impurities in Drug Substances (July 2009) 
515   
516  Changes to an Approved NDA or ANDA (April 2004) 
517   
518  Changes to an Approved Application for Specified Biotechnology and Specified Synthetic 
519  Biological Products (July 1997) 
520   
521  Changes to an Approved NDA or ANDA; Specifications  Use of Enforcement Discretion for 
522  Compendial Changes (November 2004) 
523   
524  Content and Format of Investigational New Drug Applications (INDs) for Phase 1 Studies of 
525  Drugs, Including Well-Characterized, Therapeutic, Biotechnology-derived Products (November 
526  1995) 
527   
528  IND Meetings for Human Drugs and Biologics, Chemistry Manufacturing and Controls 
529  Information (May 2001) 
530   
531  INDs for Phase 2 and 3 Studies of Drugs, Including Specified Therapeutic Biotechnology-
532  Derived Products (February 1999) 
533   
534  Investigating Out of Specification (OOS) Test Results for Pharmaceutical Production (October 
535  2006) 
536   
537  Process Validation: General Principles and Practices (January 2011) 
538   
539  Reviewer Guidance, Validation of Chromatographic Methods (November 1994) 
540   
541  Submission of Chemistry, Manufacturing, and Controls Information for Synthetic Peptide 
542  Substances (November 1994) 
543   
                                                           
27 Draft guidances have been included for completeness only.  As draft documents, they are not intended to be 
implemented until published in final form. We update guidances periodically. To make sure you have the most 
recent version of a guidance, check the FDA Drugs guidance Web page at 
http://www.fda.gov/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/default.htm.  
 
  13  Contains Nonbinding Recommendations 
544  Guidance for Industry:  International Conference on Harmonization  
545   
546  Q1A(R2) Stability Testing of New Drug Substances and Products (November 2003) 
547   
548  Q1B Stability Testing:  Photostability Testing of New Drug Substances and Products (May 
549  1997) 
550   
551  Q1C Stability Testing for New Dosage Forms (May 1997) 
552   
553  Q2(R1) Validation of Analytical Procedures: Text and Methodology (March 1995, May 1997) 
554   
555  Q3A(R2) Impurities in New Drug Substances (June 2008) 
556   
557  Q3B(R2) Impurities in New Drug Products (August 2006) 
558   
559  Q3C Impurities:  Residual Solvents (December 1997)  
560   
561  Q3C Tables and List (February 2012) 
562   
563  Q5C Quality of Biotechnological Products:  Stability Testing of Biotechnological/Biological 
564  Products (July 1996) 
565   
566  Q6A Specifications:  Test Procedures and Acceptance Criteria for New Drug Substances and 
567  New Drug Products:  Chemical Substances (December 2000) 
568   
569  Q6B Specifications:  Test Procedures and Acceptance Criteria for Biotechnological/Biological 
570  Products (August 1999) 
571   
572  Q7 Good Manufacturing Practice Guide for Active Pharmaceutical Ingredients  
573  (August 2001) 
574   
575  United States Pharmacopeia/National Formulary 
576   
577  General Chapter <621> Chromatography 
578   
579  General Chapter <1010> Analytical Data  Interpretation and Treatment 
580   
581  General Chapter <1224> Transfer of Analytical Procedures 
582   
583  General Chapter <1225> Validation of Compendial Procedures 
584   
585  General Chapter <1226> Verification of Compendial Procedures 
586   
587  General Notices and Requirements, Applying to Standards, Tests, Assays, and Other 
588  Specifications of the United States Pharmacopeia: 7. Test Results 
589   
  14  Contains Nonbinding Recommendations 
590  Interpretation and Treatment of Analytical Data; USP Pharmacopeial Forum, United States 
591  Pharmacopeial Convention, Inc., Rockville MD:  1994, Volume 24, Number 5, pp. 7051 - 7056 
592   
593  Other 
594   
595  ASTM Standard, E29 - 2008 Standard Practice for Using Significant Digits in Test Data to 
596  Determine Conformance with Specifications, ASTM International, West Conshohocken, PA, 
597  (www.astm.org). 
598   
599  ASTM E1488  Standard Guide for Statistical Procedures to use in Developing and Applying 
600  Test Methods, ASTM International, West Conshohocken, PA, (www.astm.org). 
601   
602  ASTM E2782  Standard Guide for Measurement Systems Analysis (MSA), ASTM 
603  International, West Conshohocken, PA, (www.astm.org). 
604   
605  ASTM Standard, E2935  2013 Standard Practice for Conducting Equivalence Testing in 
606  Laboratory Applications, ASTM International, West Conshohocken, PA, (www.astm.org).  
607   
608  J.N. Miller and Miller, J.C., 2010, Statistics and Chemometrics for Analytical Chemistry, 6th 
609  edition, Pearson Education Canada. 
610   
611  Saunders, B.D. and R.G. Trapp, 2004, Basic and Clinical Biostatistics, 4th edition, Lange 
612  Medical Books/McGraw Hill. 
613   
614   
615   
616   
617   
  15