Development of an ELISA-kit for a Rapid Identification of Pathogenic Bacteria

为了诊断细菌感染，有必要确定病原体。临床微生物实验室常规使用的方法是基于分离和检测分离的致病菌的生化特性。然而，这些方法通常需要20-30小时，并且在某些情况下超过72小时，因此可用的信息太晚而不能被适当地考虑用于感染的适当治疗。本研究的目的是建立一种快速、灵敏的方法来鉴定病原菌，该方法基于已成功开发的用于检测细菌蛋白毒素的珠-ELISA。因此，即使有必要将样品稀释一千倍，每ml可检测10^3个细菌的ELISA也足以检测目标细菌。flexneri，S. boydii，S.宋内氏弧菌、霍乱弧菌、副溶血弧菌和空肠弯曲杆菌制备，并且通过用适当处理的抗体进行凝集试验来确认每种抗体仅与同源细菌反应的特异性。通过使用这些特异性抗体，在检查各种反应条件后，开发了珠-ELISA系统。用上面列出的各种细菌制备了成功的ELISA试剂盒，特别是用于鉴定空肠弯曲菌的试剂盒将在临床微生物学实验室中进行测试。

To diagnose bacterial infections, it is necessary to identify the causative agent. The methods routinely applied at clinical microbiology laboratory are based on isolation and examination of biochemical characters of the isolated causative bacteria. However, these methods usually require 20-30 hours, and in some instances more than 72 hours, thus the informations available are too late to be properly considered for a proper therapy of the infections. The present investigation is aimed to develop a rapid, sensitive method to identify pathogenic bacteria based on the bead-ELISA that has successfully been developed for detecting bacterial protein toxins.In diarrheal stools, causative bacteria are usually included in the order of 10^6 bacteria per ml. Thus, even if it is necessary to dilute the specimen a thousand folds, an ELISA that can detect 10^3 bacteria per ml is sufficient for the detection of the objective bacteria.Polyclonal antibodies against whole cell preparations of Shigella dysenteriae, S. flexneri, S. boydii, S. sonnei, Vibrio cholerae, V. parpahamemolyticus and Campylobacter jejuni were prepared and the specificities of each antibody to react only the homologous bacteria were confirmed by an agglutination test with appropriately treated antibodies. By using these specific antibodies, a bead-ELISA system was developed after examining various conditions of the reaction. A successful ELISA-kit was prepared with various bacteria listed above, and especially a kit to identify Campylobacter jejuni is to be tested in clinical microbiology laboratories.