Kaslow H R, Platler B W, Blumberg D A, Cherry J D
Department of Physiology and Biophysics, University of Southern California School of Medicine, Los Angeles 90033.
J Clin Microbiol. 1992 Jun;30(6):1380-7. doi: 10.1128/jcm.30.6.1380-1387.1992.
Bordetella pertussis produces a protein virulence factor termed pertussis toxin. Many candidate pertussis vaccines are based on the rationale that an immune response that neutralizes the virulence activities of this toxin, which are thought to arise from its catalytic ADP-ribosyltransferase activity, would be beneficial. The report describes two methods that quantify the inhibition of this activity by human serum. One, termed a direct assay, involves an initial incubation of toxin with serum, a second incubation that activates the toxin, and a third incubation that measures the ADP-ribosyltransferase activity of the mixture. The other assay, termed a plate assay, involves immobilization of the toxin, exposure of the immobilized toxin to serum and washing of the plate, and then activation and assay of the toxin's ADP-ribosyltransferase activity. The plate assay may be more selective than the direct assay in terms of identifying antibodies that neutralize the toxin in vivo. Sera from controls, selected patients presenting with cough, and vaccinated infants were first analyzed by the direct assay. In contrast to sera from controls, sera from several of the patients and vaccinated infants strongly inhibited activity. Dose-response curves of inhibition were determined for samples from three vaccinated infants by both the direct and plate assays. One of the samples had a dose-response curve of a different shape and thus differed not only in titer but also in functional characteristics. A comparison of inhibition of ADP-ribosyltransferase activity and neutralization in a CHO cell assay indicated that there was incomplete agreement between the two assays. Taken together, these results indicate that measurement of inhibition of ADP-ribosyltransferase activity by human serum is practical and may be useful in the evaluation of responses to pertussis vaccines.
百日咳博德特氏菌产生一种名为百日咳毒素的蛋白质毒力因子。许多候选百日咳疫苗的设计依据是，能中和该毒素毒力活性的免疫反应会有益处，而这种毒力活性被认为源自其催化性ADP核糖基转移酶活性。该报告描述了两种定量人血清对这种活性抑制作用的方法。一种称为直接测定法，包括毒素与血清的初次孵育、激活毒素的第二次孵育以及测量混合物ADP核糖基转移酶活性的第三次孵育。另一种测定法称为平板测定法，包括固定毒素、将固定的毒素暴露于血清并冲洗平板，然后激活并测定毒素的ADP核糖基转移酶活性。就识别体内中和毒素的抗体而言，平板测定法可能比直接测定法更具选择性。首先通过直接测定法分析了来自对照组、出现咳嗽症状的选定患者以及接种疫苗婴儿的血清。与对照组血清不同，一些患者和接种疫苗婴儿的血清强烈抑制活性。通过直接测定法和平板测定法确定了来自三名接种疫苗婴儿样本的抑制剂量反应曲线。其中一个样本的剂量反应曲线形状不同，因此不仅效价不同，功能特性也不同。在CHO细胞测定中对ADP核糖基转移酶活性抑制作用和中和作用的比较表明，这两种测定法之间不完全一致。综上所述，这些结果表明，测量人血清对ADP核糖基转移酶活性的抑制作用是可行的，可能有助于评估对百日咳疫苗的反应。
