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An improved monocyte activation test using cryopreserved pooled human mononuclear cells.

Shabnam Solati1, Lucien Aarden1, Sacha Zeerleder2

  • 1Department of Immunopathology, Sanquin Research and Landsteiner Laboratory AMC, University of Amsterdam, Amsterdam, the Netherlands.

Innate Immunity
|April 25, 2015
PubMed
Summary

The monocyte activation test (MAT) using frozen pooled cells offers a reliable alternative to the rabbit pyrogen test. This method accurately detects various pyrogens in injectable drugs, ensuring product safety.

Keywords:
Cryopreserved pooled human mononuclear cellsinterleukin-6monocyte-activation testpyrogens

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Area of Science:

  • Biotechnology
  • Immunology
  • Pharmaceutical Science

Background:

  • The rabbit pyrogen test is the current standard for detecting pyrogens in injectable drugs but has limitations.
  • The monocyte activation test (MAT) is a promising alternative, utilizing human monocytes to detect pyrogenic cytokines.
  • Challenges with the MAT include donor variability and the need for fresh cells, hindering laboratory accessibility.

Purpose of the Study:

  • To establish a reproducible and accessible monocyte activation test (MAT) using frozen pooled human mononuclear cells (MNCs).
  • To demonstrate the sensitivity and specificity of the MAT with frozen pooled MNCs for detecting endotoxin and non-endotoxin pyrogens.
  • To validate the use of frozen pooled MNCs as a practical solution for routine pyrogen testing in pharmaceutical quality control.

Main Methods:

  • Developing a MAT protocol using thawed, pooled human MNCs co-cultured overnight with test substances.
  • Measuring interleukin-6 (IL-6) production as the primary readout for pyrogenic activity.
  • Comparing the sensitivity and reproducibility of the MAT using frozen pooled MNCs against fresh MNCs and different pyrogen types.

Main Results:

  • Frozen pooled MNCs demonstrated comparable sensitivity to fresh MNCs in the MAT.
  • The MAT performed with different batches of frozen pooled MNCs exhibited excellent reproducibility.
  • The test successfully detected IL-6 induction by various non-endotoxin pyrogens, confirming its broad applicability.

Conclusions:

  • The monocyte activation test (MAT) utilizing frozen pooled MNCs is a highly sensitive, specific, and reproducible pyrogen detection method.
  • This approach overcomes the limitations of donor variation and fresh cell accessibility associated with traditional MAT.
  • The MAT with frozen pooled MNCs is suitable for detecting and quantifying both endotoxin and non-endotoxin pyrogenic contaminants in parenteral pharmaceuticals.