Monocyte activation test as a pyrogen testing method

Abstract

Pyrogens are fever-causing substances that can cause serious symptoms when they enter the human body and bloodstream. The most prominent pyrogens are the lipopolysaccharides of the outer cell membranes of gram-negative bacteria. Lipopolysaccharides are endotoxins, but pyrogens can also be non-endotoxins, such as parasites or components of viruses and gram-positive bacteria.

Parenteral drug products and medical devices must be tested for pyrogens before entering the market. The most traditional way of testing pyrogenicity is the rabbit pyrogen test. The weaknesses of the test, in addition to the use of experimental animals, are its low sensitivity and non-quantitative nature. Another frequently used test is the limulus amebocyte lysate test, which is an in vitro test, but its major weakness is its ability to detect only endotoxins.

The monocyte activation test is a newer method of testing pyrogens than the test methods mentioned above. It was included in the European Pharmacopoeia in 2010, and in 2016 it was recommended to replace tests on rabbits with the monocyte activation test wherever possible. According to the United States Pharmacopeia, a validated monocyte activation test can be used instead of the traditional test methods when expedient.

The version of the monocyte activation test used in the study utilizes human peripheral blood mononuclear cells and the cytokine interleukin 6 they produce. Monocytic cells have Toll-like receptors that recognize pyrogens, causing the characteristic signaling pathways to be activated. Ultimately, the pathways lead to the activation of specific gene expressions that induce the production of inflammatory cytokines. The possibly produced interleukin 6 can then be measured with a specific enzyme-linked immunosorbent assay.