Monocyte activation test (MAT) reliably detects pyrogens in parenteral formulations of human serum albumin

Abstract

Disadvantages of the regulatory pyrogen test to assure safety of the end-product Human Serum Albumin (HSA) for parenteral use call for the implementation of an alternative test. In the current study, 16 HSA batches were assayed for pyrogens in parallel with the Rabbit Pyrogen Test, conventional and endotoxin-specific LAL assay and monocyte activation test (MAT). It was found that all HSA batches were contaminated with (1,3)-β-glucans, which interfere with the conventional LAL. Endotoxin-specific LAL was not suitable to test HSA due to unacceptable endotoxin recovery. Experiments combining polymyxin B and MAT demonstrated that pyrogenic batches were mainly contaminated with endotoxins. However, endotoxin-specific LAL failed to detect one of them. The contaminating (1,3)-β-glucans enhanced the MAT/IL-6 response to endotoxin, but not that of MAT/IL-1β. The endotoxin equivalent concentrations obtained using the IL-6 readout were usually higher than those using IL-1β, probably owing to the direct induction of IL-6 release from monocytes by (1,3)-β-glucans. The MAT correlates with the rabbit pyrogen test, providing a higher safety level for pyrogenicity testing of HSA and probably other therapeutic proteins.