Enhancing the Accuracy of Semi-quantitative Endotoxin Evaluation in Biopharmaceuticals and Vaccines Biopharmaceuticals and Vaccines R D

Introduction: The detection of endotoxins is crucial in the research and development of new drugs and vaccines, as it ensures the safety of these products. The quantitative Limulus Amebocyte Lysate (LAL) endotoxin tests provide sensitive and accurate results. Quantitative endotoxin tests may be substituted by LAL gel clot test with enough narrow dilution range as a semi-quantitative method. However, the accuracy and reliability of the assay can be affected by the dilution factor used. Methods: The endotoxin concentration of different samples of a bench top purification process of recombinant streptokinase, including inclusion body, washed inclusion body, semi-purified and purified streptokinase was determined by semi-quantitative LAL gel clot and quantitative LAL chromogenic test and the effects of narrow-downing the dilution range of the samples on the accuracy of the results was evaluated. Results: The statistical analysis revealed that performing duplicate LAL gel clot tests and consecutively narrowing the dilution range of the sample until at least a positive and a negative results were seen, offers a good estimation of the endotoxin concentration. The relative errors of these results were less than 12%, compared to accurate results of quantitative methods. However, conducting gel clot test at the wide dilution range for the inclusion body samples resulted in approximately 200% overestimation. Conclusion: the results suggest that the semi-quantitative LAL gel clot test with a narrow dilution range can be a valuable tool for relatively accurate estimation of endotoxin in biopharmaceutical products including vaccines.