Detection of endotoxins in human blood and plasma. An improved in-vitro pyrogen test

We describe an improved in-vitro procedure for detection of endotoxin in human blood and plasma by use of Limulus amoebocyte lysate. Increasing concentrations of Escherichia coli endotoxin added to a constant amount of the lysate cause a proportional increase in protein precipitated by the endotoxin. By measuring the amount of protein precipitated, it was possible to determine the equivalent E. coli endotoxin concentration in unknown samples, when samples were run with E. coli endotoxin standards and negative controls. The E. coli endotoxin, present in human whole blood and platelet-rich plasma, failed to react with the lysate. However, the concentration of endotoxin in whole blood and platelet-rich plasma could be measured with this Limulus test after lysing the platelets to release the endotoxin and subsequently removing the inhibitory proteins by chloroform precipitation. With this procedure it was possible accurately and repeatedly to determine E. coli equivalent endotoxin concentrations as low as 195 ng per liter of whole blood or 49 ng per liter of platelet-rich plasma.