비탁법에 의한 환경 중 엔도톡신의 정량분석방법에서 반응액 주입시간의 차이에 의한 영향 및 보정방법에 관한 연구

Abstract

Workers can be exposed to endotoxins released by Gram-negative bacteria, which can cause a response in humans ranging from mild fever to shock or even death, from their work environments such as cotton processing, animal handling and farming, grain handling and use of metalworking fluids. An analytical method has been developed for endotoxin based on the reaction of LAL with LPS, that is known to be easy, fast, and sensitive. Quantitative LAL methods based on turbidimetric assays are commonly used for workplace samples. It takes more than 90 seconds(up to 180 seconds upon the dexterity) to inject reaction reagents to all cells of samples in the 96 well micoplate during preparing samples for analysis based on turbidimetric assays. These injection time intervals between samples resulted in actual reaction duration before reading the optical density by the instrument, and resulted in erroneous quantitative results. This study was conducted to estimate the effects of injection time interval to the quantitative results and to develop a correction method. The results of this study were as in the follows. 1) Time delay due to injection time intervals between cells in the 96 well microplates resulted in decrease of endotoxin concentration significantly. It was found that the endotoxin concentration decreases as time delay increases. 2) It was shown that these effects were greater at a high concentration levels(10 EU/ml) compared to a relative low concentration levels(1 EU/ml). 3) It was able to demonstrate that these errors could be reasonably corrected by applying a correction equation derived fTom the experiments as shown on the below. Y=TRi-Ti(Ci-1) here, Y: corrected reaction time(for use to detennine OD) TRi: reaction time of ith column read by the instrument to reach the predetennined OD Ci: Number of i^(th) column T=((Total elapsed time for injection to 12 column)-T_(1))/(C_(i)-1)