Effect of Human Serum Albumin on Endotoxin Scattering Photometry

Abstract

Laser scattering photometry (ESP) is a newly developed plasma endotoxin assay method using horseshoe crab amebocyte lysate (AL) that recognizes small particles produced by polymerization of coagulin under the stirring conditions at 1000rpm. We elucidated the effect of human serum album (HSA) in the ESP method. AL was dissolved with 630μL of the specimen and a 200-μL aliquot was used for ESP; this conventional protocol was regarded as the ESP630 method. The ESP210 method was also used, i. e. AL was dissolved with 210μL of the specimen and a 200-μL aliquot was used for ESP. Water induced the agglutination, and HSA prolonged the agglutination time depending on its concentration especially in the ESP630 method. The water-induced agglutination was not inhibited by the addition of anti-factor C monoclonal antibody, and amidinophenyl benzoate hydrochloride, used as a clotting enzyme inhibitor, intensively inhibited the water-induced agglutination. Therefore, the water-induced agglutination was suggested to be a false-positive reaction to non-specific activation of the clotting enzyme. The HSA-induced prolongation of the reaction in the national health insurance-covered turbidimetric kinetic assay was not observed. HSA or plasma protein seemed to affect the result, especially in the ESP630 method, and a non-specific reaction was found to occur in the ESP methods.