What does cleaning validation mean in practice?
In practice, cleaning validation answers the question of whether the described cleaning process—manual or washer-disinfector-assisted—reproduces sufficiently high levels of cleanliness in a reproducible manner. This includes, in particular:
Cleansing markers: protein, hemoglobin, TOC
- Protein is considered a sensitive marker for organic clinical contamination (blood, tissue).
- Hemoglobin is specific for blood-containing contaminants and is recommended as an additional marker in many standards.
- TOC (Total Organic Carbon) captures the total content of organic residues and ideally complements protein/hemoglobin — especially in implants and complex geometries.
Requirements for implants – ISO 19227
Additional cleanliness requirements apply to orthopedic implants:
- quantitative TOC assessment
- Particulate cleanliness according to AAMI TIR42
- chemical and biological purity tests
- If necessary, cytotoxicity or endotoxin tests as follow-up investigations.
ISO 19227 is therefore a key certification for implant manufacturers.
Supplementary microbiological and toxicological tests are also required.
If cleanliness or particle analyses show abnormal results, bioburden testing according to ISO 11737-1 and cytotoxicological assessments according to ISO 10993-5 may be necessary.
These follow-up tests help to assess whether residual contamination is microbiological or chemical-toxicological in nature and support the overall assessment according to EN ISO 17664 and MDR.
Supplementary microbiological and toxicological assessments:
If TOC, protein, hemoglobin, or particle analyses show abnormal results, bioburden testing according to ISO 11737-1 and cytotoxicological investigations according to ISO 10993-5 may be required. These follow-up tests help to clarify whether remaining residues are microbiologically or chemically-toxicologically relevant and support the overall assessment according to ISO 17664.
Normative testing methods
- ISO 15883-5 : defined cleaning test methods, artificial soiling
- ANSI/AAMI ST98 : Structured Validation for Cleaning Processes (US Market)
- ASTM F3208-20 : Standardized Soiling Kits for Worst-Case Loads
- USP 643 : Analytical specifications for TOC measurements
- AAMI TIR42 : Measurement and evaluation of particles
HygCen Germany conducts these tests in accordance with standards and evaluates only the reproducible effectiveness — without process consulting or optimization.
Standard criteria – at a glance
| Standard criterion | Requirements / Content | HygCen test |
|---|---|---|
| Cleaning marker | Protein, hemoglobin, TOC | quantitative residue analysis |
| Implant purity | ISO 19227: TOC, particles, chemical residues | implant-specific testing programs |
| Cleaning methods | ISO 15883-5, ST98, ASTM F3208-20 | Implementation of defined soiling and testing methods |
| Particle test | AAMI TIR42 | quantitative and qualitative particle assessment |
| Endotoxins | ISO 10993-11 (LAL) | optional for critical products |
| microbiology | ISO 11737-1/-2 | supplementary microbiological considerations |
| Bioburden | ISO 11737-1: Quantitative assessment of residual microbial contamination | Microbiological follow-up testing in case of conspicuous cleanliness parameters |
| Cytotoxicity | ISO 10993-5: Cellular compatibility of chemical residues | supplementary assessment for chemically/toxicologically relevant residues |
| documentation | QES signed, auditable | MDR-compliant test certificates |
This is how we review your cleaning processes – step by step
Analysis of the process described by the manufacturer/operator.
We consider:
- Cleaning methods used (manual/washer-disinfector)
- Programs, media, exposure times, temperature profiles
- Loading types (instruments, implants, hollow bodies)
- Standard-relevant specifications (ISO 17664, ISO 15883-5, ST98, ISO 19227)
No advice, only testability and validation.
- Test planning & selection of cleaning markers
We define:
- Protein, hemoglobin or TOC — individually or in combination
- ASTM or ISO compliant soiling
- Worst-case test specimens (e.g. hollow bodies, rough surfaces, implant surfaces)
- Measurement methods & evaluation procedures
- Number of cycles & repetitions
Laboratory testing under realistic conditions.
We conduct:
- quantitative residual analysis (protein/Hb/TOC)
- Particle measurements (AAMI TIR42)
- Implant-specific purity tests (ISO 19227)
- LAL/Endotoxin (if required in the process)
- microbiological supplements if needed
All tests are DAkkS-accredited and neutral.
- Validability assessment
We assess:
- whether the process steps produce reproducible cleanliness
- whether the results meet the standard criteria
- whether the process is considered validatable in the IFU (ISO 17664)
- whether a consistent interface to RDG and sterilization will result
- QES-signed test report
- The test report contains:
- complete test description
- quantitative results
- Standard reference & evaluation
- Statement on validation
- no process or consulting components
Why HygCen?
- Independent, DAkkS-accredited testing laboratory with 30 years of experience
- Neutral cleaning and cleanliness assessments without process consulting
- Coverage of all relevant standards (ISO 19227, 15883 5, ST98, F3208, USP 643)
- QES-signed reports for MDR, IFU validation & audits
- Can be combined with RDG, sterilization, packaging, microbiology and end-of-life validation
Protein is considered a sensitive indicator for organic clinical soiling and is defined as a key marker in ISO 15883-5 and ANSI/AAMI ST98. Hemoglobin serves as a specific marker for blood-containing soiling and is often used in parallel when blood is considered a realistic worst-case contamination. TOC, in turn, reflects the total organic residue content and is particularly suitable for critical products, implants, and complex structures. In combination, these markers provide a very comprehensive picture of cleaning performance—from specific organic residues to sum parameters.
ISO 19227 applies to orthopedic implants and similar products where chemical and particulate cleanliness is critical. The standard requires, among other things, quantitative TOC limits, particle assessment according to AAMI TIR42, identification of potential residues, and traceable documentation of the entire cleaning process. HygCen only verifies compliance with these requirements; process design and the selection of suitable cleaning methods remain the responsibility of the manufacturer or external consultants. The results support the technical documentation and serve as a basis for risk-based decisions.
In the case of borderline results, HygCen Germany conducts a purely data-based assessment – without process consulting or optimization recommendations. If several test parameters (e.g., protein and hemoglobin) are close to acceptance limits or show variable results, we document this precisely in the test report. The operator or manufacturer must then consult with process or regulatory advisors to review possible adjustments (e.g., loading, chemicals, exposure time). HygCen Germany conducts repeat tests upon request but does not evaluate process improvement strategies.
Many manufacturers require cleanliness certifications for both the European and US markets. ASTM F3208-20 provides standardized soiling methods, ST98 defines structured cleaning validation procedures, and USP 643 specifies analytical criteria for TOC measurements. HygCen Germany can combine these methods in testing programs to enable international comparability. It is important to note that HygCen Germany tests these methods but does not advise on which method is strategically ideal for which market—these decisions are the responsibility of external regulatory experts.
Revalidation is always required when safety-relevant factors change: new instrument types, different process chemicals, adjustments to washer-disinfector (WD) programs, changes in water quality, or unusual test results. Furthermore, many quality management (QM) systems stipulate regular intervals. HygCen Germany documents the validatability of current processes.
Bioburden testing (ISO 11737-1) or cytotoxicity testing (ISO 10993-5) becomes relevant when quantitative cleanliness parameters such as TOC, protein, hemoglobin, or particles (AAMI TIR42) are abnormal, or when the product is used near the implant. These follow-up tests help to clarify whether any remaining residual contamination is microbiologically or chemically toxicologically relevant.
Further links
ISO 17664 - Validation of manufacturer's specifications, ISO 15883 - Waste Disposal Device Validation, ISO 17665-1 / EN 13060 / EN 285 - Sterilization Validation, End of Life - Material and Functional Durability, Bioburden & Sterility - ISO 11737-1/-2, Type Testing of Reprocessing Equipment, Packaging & sterile barrier – ISO11607-2, Reprocessing of medical devices