Cleaning Validation Protocol Format and Acceptance Criteria

In short: This is an illustrative cleaning validation protocol format for QA, QC and production staff, with example acceptance criteria for the analytical method and swab recovery. All numbers shown are examples only and must be justified and approved under your own quality system. Carry-over limits must come from health-based exposure limits (PDE/ADE), not from the numbers in this post.

Template notice: adapt section titles, numbering and every acceptance criterion to your site’s approved procedures.

Protocol format

Prepare one protocol for the selected worst-case product (see the worst-case rationale in the SOP). Number it as per your site’s protocol numbering SOP. Typical sections:

Section	What it contains
Objective	Purpose of the study
Scope	Product, equipment train, site and cleaning procedure covered
Responsibilities	Who prepares, executes, samples, analyses and approves
Training	Training of everyone involved, with records, including GMP awareness
Signature log	Specimen signatures and initials for identification
Cleaning procedure	Equipment, reference cleaning SOP and full cleaning steps, including cleaning agent, time and temperature
Sampling procedure	Visual inspection, swab preparation, swab area, sampling locations with rationale, rinse sampling if used
Analytical and microbiological methods	Reference to validated methods for residues and bioburden
Acceptance criteria	Visual, residue and microbial limits with the HBEL-based calculation
Number of runs	Justified by risk assessment
Hold times	Dirty and clean hold times being challenged
Deviations	How deviations are recorded; justification approved by QA head and plant head as per site SOP
Report	Summary of results for each equipment item after all runs are complete

Sampling

Take swab samples after final cleaning and only after the equipment passes visual inspection. Define the swab preparation, the area swabbed (for example 25 cm², site-defined), and the swab points, selecting the hardest-to-clean locations. Attach each activity record to the protocol.

Acceptance criteria

State all criteria before execution:

Equipment is visually clean.
Residue of the previous product is below the limit derived from its PDE/ADE (health-based exposure limit), converted to a per-swab or per-area value for the shared equipment train. Include the calculation and the toxicological reference.
Cleaning-agent residue is below its justified limit.
Microbial and endotoxin limits, where relevant, are met.

Visual cleanliness is an important criterion but not sufficient alone, and repeated cleaning and retesting until results pass is not acceptable (EU GMP Annex 15, clauses 10.2 and 10.6). The older 10 ppm and 1/1000 dose rules are legacy criteria that may be kept as secondary checks only.

The report is compiled after the planned number of runs. Annex 15 requires “an appropriate number of times based on a risk assessment” (clause 10.13). Three consecutive runs is the in-house practice of triplicate runs, used to show reproducibility of the cleaning results; it is a site criterion, not a number set by Annex 15.

Analytical method validation for cleaning samples

The methods used for residue and bioburden testing must be validated, with an appropriate detection and quantification limit where necessary (EU GMP Annex 15, section 9.1). ICH Q2(R2) sets the validation characteristics, but it does not prescribe numeric limits for each, so the criteria below are examples only, commonly seen in industry protocols. Each site must set and justify its own criteria.

Parameter	What is checked	Example criterion (illustrative)
Limit of quantification (LOQ) precision	Repeatability of triplicate injections at LOQ	RSD of peak area not more than 10.0%
Limit of detection (LOD)	Lowest concentration at which the analyte is detected	Often estimated from signal-to-noise (about 3:1) and confirmed by injections
Linearity	Response proportional to concentration, from LOQ up to about 150% of the acceptance limit	Correlation coefficient not less than (NLT) 0.990; plot visually linear. Also review y-intercept and residuals
Precision at LOQ	Six replicate injections of the standard, and six preparations of the spiked sample	RSD of area not more than 10%; RSD of recovery not more than 10%
Accuracy at LOQ	Recovery from placebo spiked at LOQ	80.0% to 120.0%
Solution stability	Standard and sample solutions held at room temperature	Change from initial not more than 10% over the stated interval
Recovery from spiked swab	Known amount spiked and swabbed	Not less than 80%
Recovery from product-contact surface (for example SS plate)	Efficiency of the swab technique on the actual material	Mean not less than 70%

Correction note: an earlier version of this post said the linearity correlation coefficient “should be less than 0.990”. That was wrong. The correlation coefficient must be not less than the stated value, so values closer to 1.000 pass.

Recovery factors from the surface study should be applied when evaluating cleaning validation results. Annex 15 clause 10.12 expects recovery to be shown for every product-contact material sampled, with each sampling method used. Different sites also use lower recovery bands with justification.

Common myths and facts

Myth 1: “You cannot validate manual cleaning”

Manual cleaning can be validated. It varies more than automated cleaning, so it needs a very detailed procedure, trained operators and adequate cleaning steps. Annex 15 (clause 10.15) says the effectiveness of manual cleaning should be confirmed at a justified frequency.

Myth 2: “Recovery at different spike levels must be linear”

Swabbing is a manual technique, and recovery results vary. A strictly linear recovery response across spike levels is not a general requirement. What matters is that recovery is reproducible, acceptable and used to evaluate results.

Myth 3: “Any detectable residue means failure”

Methods are sensitive enough that some residue is often detectable. A result below the justified, health-based acceptance limit, with no effect on product quality, is acceptable.

Myth 4: “HPLC must always be used instead of TOC”

Annex 15 (clause 10.6.2) allows representative parameters such as TOC and conductivity where testing for a specific residue is not feasible. A validated TOC method, suited to the residue, can be acceptable without duplicate testing by HPLC. Justify its suitability for the specific residue and the limit.

Myth 5: “Rinse results must correlate with swab results”

They are different samples. A swab samples a small, defined, hardest-to-clean spot, while a rinse samples a larger area, including parts that cannot be reached. The FDA inspection guide notes that direct surface sampling is preferred and that rinse samples can miss insoluble or trapped residue, so each is justified for its own purpose rather than being forced to match.

Frequently asked questions

Should the correlation coefficient be below or above 0.990?

Not less than the stated value, commonly 0.990 or higher, set by the site. A value below the limit means poor linearity.

Who sets the cleaning limits?

A qualified toxicologist derives the PDE/ADE. QA converts it into a per-swab or rinse limit for the protocol.

Do I need to validate swab recovery?

Yes. Recovery must be shown for each product-contact material and each sampling method (Annex 15, clause 10.12).

Do I need a microbiological limit?

Annex 15 clause 10.7 says microbial and endotoxin risk should be considered when developing protocols. Set limits according to the product risk.

References

European Commission, EudraLex Volume 4, EU GMP Annex 15: Qualification and Validation (Ares(2015)1380025, 30 March 2015; deadline for coming into operation 1 October 2015), sections 9 and 10.
ICH, Q2(R2): Validation of Analytical Procedures (adopted November 2023).
European Medicines Agency, Guideline on setting health based exposure limits for use in risk identification in the manufacture of different medicinal products in shared facilities, EMA/CHMP/CVMP/SWP/169430/2012 (2014).
US FDA, Guide to Inspections: Validation of Cleaning Processes (7/93).
PIC/S, PI 046-1 (1 July 2018), Guideline on setting health based exposure limits.