Pyrogen testing
Testing for pyrogens is a critical step in ensuring medical products administered parenterally, such as pharmaceuticals or biopharmaceuticals safety. It is part of the mandatory release tests to avoid life-threatening fever reactions induced by pyrogenic substances. The monocyte activation test (MAT) can detect both endotoxin and non-endotoxin pyrogens in one in vitro test.
Since July 2025, the MAT has been the method of choice according to European Pharmacopoeia for endotoxin and non-endotoxin detection, replacing the Rabbit Pyrogen Test (RPT).
Products
Monocyte Activation Test (MAT)
Used to detect both endotoxins and non-endotoxin pyrogens in parenteral products, such as pharmaceuticals and biopharmaceuticals, the MAT gives an in vitro alternative to conventional animal testing in accordance with regulatory guidelines.
The Rabbit Pyrogen Test and the Limulus Amebocyte Lysate (LAL) test are broadly used for pyrogen detection. Both methods use animals and show some limitations. The rabbit pyrogen test shows a lack of robustness as an animal reaction can differ greatly from a human reaction. In the LAL test, only endotoxins are detected causing a safety risk by ignoring non-endotoxin pyrogens that could be present in the tested sample.
To overcome these limitations, the Monocyte-Activation Test (MAT) was introduced in the European Pharmacopoeia in 2010 as a compendial method to replace the Rabbit Pyrogen Test (EP Chapter 2.6.30) and mentioned in FDA guidance for industry.
Please Note: The European Pharmacopoeia Commission took the decision to put an end to the rabbit pyrogen test in accordance with the 3Rs principle considering that the MAT is the best alternative option. From July 2025, the Rabbit Pyrogen Testing (RPT) is forbidden in Europe and the general chapter 2.6.8 on Rabbit Pyrogen Testing has been suppressed.
PyroMAT® in vitro test to detect endotoxin and non-endotoxin pyrogens
The PyroMAT® system is based on the Mono-Mac-6 cell line and IL-6 read-out. It offers all the advantages of the monocyte activation test combined with the benefits of using a cell line.
- Detection of a broad range of pyrogens: patient safety is ensured if the full range of pyrogens are tested. Like the rabbit pyrogen test (RPT), MAT is effective for both endotoxin and non-endotoxin pyrogen detection.
- Extension of the range of products that can be tested: the most frequently applied methods, RPT, Bacterial Endotoxin Test (BET) or LAL, are limited in the product types they are able to test. The MAT offers more flexibility regarding its applications.
- In vitro assay that mimics the human immune reaction: for a robust predictive model that reduces animal consumption.
- Compliance with international regulations and guidelines: in line with ethical trends of industry and regulatory authorities to decrease the use of animal-based testing.
- Standardized reactivity and high sensitivity (0.05 EU/mL). Convenience of a ready-to-use cell line reduces laborious lab work and avoids the need for a cell culture laboratory.
- Qualified cells: in addition to being cited in the international validation of MAT, Mono-Mac-6 cells are qualified for the expression of all surface Toll-Like Receptors (TLRs) to ensure the detection of a broad range of pyrogens.
Non-endotoxin pyrogen positive controls
The European Pharmacopeia Chapter 2.6.30 (Monocyte Activation Test) "the preparatory testing is to include validation of the test system using at least 2 non-endotoxin ligands for toll-like receptors, e.g. peptidoglycans, lipoteichoic acids, synthetic bacterial lipoproteins, flagellin and crude bacterial whole cell extract, at least 1 of which is to be spiked into the preparation being examined. The choice of non-endotoxin pyrogens used should reflect the most likely contaminants(s) of the preparation being examined.
To respond to this requirement, we are providing an extended range of positive controls:
- Reflecting several types of contaminants usually found in pharmaceutical production processes (Gram-negative and Gram-positive bacteria, viruses and mycoplasma).
- Targeting several monocytic toll-like receptors (TLR)
We will help you to shift from Animal-based testing to in vitro testing
Feasibility study, method development, validation and training services to support your pyrogen testing implementation, discuss with our Pyrogen Testing Experts.
Related resources
- Brochure: PyroMAT® system
Used to detect a broad range of pyrogens in parenteral products such as pharmaceuticals, biopharmaceuticals or medical devices, the MAT gives an in vitro alternative to conventional animal testing in accordance with regulatory guidelines.
- Datasheet: Non-endotoxin pyrogen positive controls
Meet regulatory requirements for Monocyte Activation Test (MAT) with our non-endotoxin pyrogen controls designed for the PyroMAT® system
