DEVELOPMENT OF A COMPETITIVE ENZYME-LINKED IMMUNOSORBENT-ASSAY (ELISA) FOR ESCHERICHIA-COLI HEAT-STABLE ENTERO-TOXIN (STA)
DEVELOPMENT OF A COMPETITIVE ENZYME-LINKED IMMUNOSORBENT-ASSAY (ELISA) FOR ESCHERICHIA-COLI HEAT-STABLE ENTERO-TOXIN (STA)
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DOI：
10.1016/0022-1759(84)90113-3
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发表时间：
1984-01-01
影响因子：
2.2
通讯作者：
ROBERTSON, DC
中科院分区：
文献类型：
--
作者：
LOCKWOOD, DE;ROBERTSON, DC
开发了一种灵敏的竞争性酶联免疫吸附测定（ELISA）来检测产肠毒素大肠杆菌（ETEC）培养上清液中的低分子量热稳定肠毒素（STa）。当使用针对戊二醛偶联 STa-牛血清白蛋白 (BSA) 缀合物产生的抗血清作为检测抗体时，在溶液中的 STa 与结合到微量滴定孔上的戊二醛偶联 STa-人血清白蛋白 (HSA) 缀合物之间观察到竞争性抑制。没有观察到与使用1-乙基-3-(3-二甲基氨基丙基)碳二亚胺或辛二亚氨酸二甲酯制备的缀合物和针对每种缀合物产生的抗血清的竞争。生物素/亲和素系统提高了测定的灵敏度，使得可以在不到 4 小时内检测到 133 pg/ml 的纯化 STa。该测定用于检测和定量人、猪和牛 ETEC 分离株的培养上清液中的 STa。未观察到与不耐热肠毒素 (LT) 或仅在仔猪中具有生物活性的 ST 形式 (STb) 发生交叉反应。定量 STa ELISA 结果显示与乳鼠生物测定和先前描述的放射免疫测定具有良好的相关性 (0.87)。定量测定经过修改，将总孵育时间缩短至不到 2 小时。定性 STa ELISA 为 ETEC 临床分离株提供快速、灵敏的检测方法，并有助于对产生 STa 的 ETEC 发生率进行流行病学研究。
A sensitive competitive enzyme-linked immunosorbent assay (ELISA) was developed to detect the low MW heat-stable enterotoxin (STa) in culture supernatant fluids of enterotoxigenic E. coli (ETEC). Competitive inhibition was observed between STa in solution and a glutaraldehyde-coupled STa-human serum albumin (HSA) conjugate bound to microtiter wells when antiserum raised against a glutaraldehyde-coupled STa-bovine serum albumin (BSA) conjugate was used as detecting antibody. No competition was observed with conjugates prepared using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide or dimethyl suberimidate and antisera raised against each conjugate. A biotin/avidin system increased the sensitivity of the assay such that 133 pg/ml of purified STa can be detected in less than 4 h. The assay was used to detect and quantify STa in culture supernatant fluids from human, porcine and bovine ETEC isolates. No cross-reactivity was observed with the heat-labile enterotoxin (LT) or the form of ST with biological activity only in piglets (STb). Results from the quantitative STa ELISA showed good correlation (0.87) with the suckling mouse bioassay and a previously described radioimmunoassay. The quantitative assay was modified to reduce the total incubation time to less than 2 h. The qualitative STa ELISA provides a rapid and sensitive assay for clinical isolates of ETEC and should facilitate epidemiological studies on the incidence of STa-producing ETEC.