The University of Konstanz is developing an animal-free alternative for testing injection solutions and other injectable drugs. The newly developed test uses human blood instead of rabbits.
This day is International Rabbit Day. The so-called rabbit pyrogen test will soon be a thing of the past. For decades, the rabbit pryogen test (RPT) was the method of choice to determine whether fever-causing substances are present in injection solutions or other injectable drugs.
After the substance to be tested has been injected into the ear vein of the rabbits, which are then restrained in a small cage, their body temperature is continuously recorded rectally. An increase in body temperature indicates contamination of the substance being tested. The rabbits are used for measurement in several tests until they are euthanized. In Germany alone, 6,000 rabbits were used for this test every year.
“A great success for the Animal welfare with an alternative method from Baden-Württemberg that does not involve animal testing,” said the State Animal Welfare Commissioner Dr. Julia Stubenbord on September 10, 2024. The monocyte activation test (MAT), also known as the “Konstanz method,” works with human blood instead of rabbits and uses a human immune reaction. It was developed 25 years ago, internationally validated in 2005, and incorporated into the European Pharmacopoeia The “Konstanz Method” was developed by the working group around Prof. Thomas Hartung and Prof. Albrecht Wendel at Doerenkamp-Zbinden Chair for alternative methods in Konstanz.
Alternative test with human blood
A sample of the substance to be tested is brought into contact with human blood and cultivated in an incubator at 37 degrees Celsius. If the sample is contaminated with pyrogens, a defense reaction is triggered. White blood cells, monocytes, act as indicators. No fever is triggered as in rabbits, but rather certain inflammatory messengers (cytokines) are formed in the blood. This messenger can then be made visible with a color reaction. This proves the contamination with the harmful pyrogens.
In June 2024, the European Commission a revised text in the European Pharmacopoeia, from which the rabbit pyrogen test was completely deleted. The changes will come into force on July 1, 2025 with the publication of the new European Pharmacopoeia.
What is shocking is the fact that an alternative non-animal testing method, the MAT test, was developed 30 years ago, but rabbit pyrogen tests were still carried out. Once the rabbit pyrogen test is removed, drug manufacturers will have to use another suitable method to test the pyrogenicity of a product. Unfortunately, there is another method of testing that uses the blue blood of horseshoe crabs. This method is also superfluous and could be replaced with the MAT.
