What Is Endotoxin?
The endotoxin test measures the quantity of lipopolysaccharide (endotoxin) originating from the cell wall of Gram-negative bacteria. Endotoxin is heat-resistant: sterilisation kills the bacterium but does not destroy the endotoxin. A sterile product can therefore still be pyrogenic and must be tested separately.
Purpose of Endotoxin
The purpose is to show that the endotoxin level in devices that contact the body via the bloodstream, cerebrospinal fluid, or implantation is below the permitted limit.
Which Products Is Endotoxin Applied To?
- Intravenous catheters and infusion sets
- Implants
- Dialysis sets
- Surgical instruments and sets
- Devices in contact with cerebrospinal fluid
In Which Sectors Is Endotoxin Used?
Standards Related to Endotoxin
- EP 2.6.14
- USP <85>
- ANSI/AAMI ST72
- ISO 11737-3
- ISO 11607end product sterile barrier packaging test
Endotoxin Conditions
- Method
- Gel-clot, turbidimetric, or chromogenic LAL methods
- Extraction
- The product's body-contacting surfaces are extracted with endotoxin-free water
- Interference control
- It must be verified that the product does not inhibit or enhance the test
Equipment for Endotoxin
- LAL test kits and reagents
- Endotoxin-free glassware
- Microplate reader (for kinetic methods)
- Dry-heat oven (for depyrogenation)
- Temperature-controlled incubator
Sample Requirements for Endotoxin
- The final, sterilised product
- The body-contacting surface area must be known
- The number of specimens is determined according to the pharmacopoeia and batch information
- The specimen must be prepared and transported in endotoxin-free containers
Duration of Endotoxin
The initial test, including the interference (inhibition/enhancement) study, takes several days; routine tests are shorter.
Endotoxin Procedure
- 1The endotoxin limit for the product is calculated.
- 2An interference study is performed, or reference is made to an existing study.
- 3The product is extracted with endotoxin-free water.
- 4The extract is mixed with the LAL reagent.
- 5It is incubated at the defined temperature.
- 6Clot formation, turbidity, or colour development is measured, depending on the method.
- 7The endotoxin quantity is calculated from the standard curve.
Endotoxin Acceptance Criteria
The acceptance criterion is the endotoxin limit calculated for the product. This limit is calculated using the formula defined in the pharmacopoeia, based on the device's route of contact and patient weight; the limit is markedly lower for devices in contact with cerebrospinal fluid.
Evaluation of Results
The result is compared with the calculated limit. If the interference study is not valid, the result cannot be interpreted; where the product suppresses the LAL reaction, a dilution factor must be determined.
Endotoxin Report
- Applied pharmacopoeia and method
- Specimen description, batch and surface area
- Calculated endotoxin limit
- Interference-study result and dilution factor
- Measured endotoxin value (EU/device or EU/mL)
- Comparison with the limit
Accreditation
EUROLAB holds internationally recognised accreditation obtained from accreditation bodies party to the IAF and ILAC multilateral recognition arrangements, under ISO/IEC 17025 for laboratory testing, ISO/IEC 17020 for inspection and surveillance, and ISO/IEC 17021-1 and ISO/IEC 17065 for system and product certification. This chain ensures the reports and certificates issued are accepted worldwide.
Whether this test can be offered within accredited scope depends on your product and the standard applied; since accreditation is method-based, the scope is confirmed in writing at the quotation stage.
Scope and authorisationsRelated Regulations
- MDR (EU) 2017/745
- Pharmacopoeia requirements
Medikal Cihaz Testleri — What Does This Testing Field Cover?
The section below describes the field that the Endotoxin test belongs to; the conditions and acceptance criteria of this particular test are given in the sections above. The scope specific to your product is confirmed in writing at the quotation stage.
Medical device testing shows that a device is safe for the patient and the user and maintains its intended performance. In this family safety is not only about avoiding harm; a device failing to perform its expected function is also treated as a risk. A monitor missing an alarm or a pump delivering the wrong dose is directly a patient safety event.
At the centre of the field is RISK MANAGEMENT, and this is what sets medical apart from other sectors. The risk management process run under ISO 14971 also determines which tests are required. The test list is not copied from a table of standards; it is derived and justified from the device's intended use, the nature and duration of patient contact, the use environment and foreseeable misuse.
Assessment consists of several separate bodies of work, each with its own set of standards. Biocompatibility runs through the ISO 10993 series, electrical device safety through the IEC 60601 series, sterilisation through ISO 11135/11137/17665, packaging through ISO 11607, software through IEC 62304 and usability through IEC 62366. The quality system (ISO 13485) is the framework that binds these together.
Which tests are required is determined by the device's CLASS and its MODE OF CONTACT WITH THE BODY. A product in brief contact with skin and an implant entering the bloodstream do not go through the same test set; the scope is set by a biological evaluation plan prepared under ISO 10993-1. That plan justifies in writing which endpoints are required and why the others are not.
Packaging and sterilisation are taken as seriously as the device itself in this field. For a sterile device the packaging is not a transport container but the barrier that maintains sterility; if the barrier is breached, the device is no longer sterile. Packaging processes are therefore validated and shelf life is supported by real-time ageing data.
For devices containing software, assessment shifts from testing towards PROCESS assurance. IEC 62304 defines which development and verification steps must be followed according to the safety class of the software; the evidence is the traceability of the development process. Testing software does not make it safe — the process record does.
The clinical side is a separate column of the technical file. Test reports show that the device is technically safe; clinical evaluation establishes that the device delivers the intended clinical benefit and that the benefit-risk balance is acceptable. Neither replaces the other.
Method groups in this field
Biocompatibility
Cytotoxicity, irritation, skin sensitisation, systemic toxicity, haemocompatibility, genotoxicity and local effects after implantation. The required endpoints are set by the ISO 10993-1 plan.
Electrical device safety
IEC 60601-1 basic safety and essential performance; EMC, alarm, usability and environmental requirements through the collateral standards.
Sterilisation validation
Validation of sterilisation processes by ethylene oxide, radiation or moist heat; bioburden and sterility assurance level studies.
Packaging
Integrity of the sterile barrier system, seal strength, distribution simulation and shelf-life studies.
Microbiological tests
Bioburden determination, sterility testing, bacterial endotoxin and, where required, carrier validation studies.
Software and usability
IEC 62304 life cycle evidence and use-error focused evaluation under IEC 62366.
Commonly confused in this field
- Cytotoxicity alone is not evidence of biocompatibility; the required set of endpoints is determined and justified according to ISO 10993-1.
- An ISO 13485 certificate does not confer CE marking; classification under the MDR, the technical file and the notified body process run separately.
- Accelerated ageing does not replace real-time ageing; a shelf life claim is supported by real-time data.
- A change of material (supplier, pigment, sterilisation method) may require the biological evaluation to be repeated; it is not a silent change.
- An assessment made without the collateral and particular standards for IEC 60601-1 is incomplete; which -1-x and -2-x standards were applied must appear in the report.
- In vitro diagnostic devices fall under the IVDR rather than the MDR, and laboratory equipment is often on the IEC 61010-1 side.
Which decisions these results support
- MDR/IVDR technical file and notified body assessment
- Completing the biological evaluation plan
- Sterilisation and packaging validation
- Determining shelf life and storage conditions
- Determining the scope of reassessment after a design change
Related Tests
Frequently Asked Questions About Endotoxin
My product is sterile — why is an endotoxin test needed?
Endotoxin is heat-resistant; although sterilisation kills the bacterium, it does not break down endotoxin, which is a cell-wall residue. A sterile product can therefore still cause a pyrogenic reaction, and endotoxin must be measured separately.
How is the endotoxin limit determined?
It is calculated using the formula defined in the pharmacopoeia, based on the device's route of contact and the permitted dose. For devices in contact with cerebrospinal fluid, the limit is much lower; the calculation must be shown in the report.