Kuo J S, Monji N, Schwalbe R S, McCoy D W
J Immunol Methods. 1981;43(1):35-47. doi: 10.1016/0022-1759(81)90034-x.
A new polyethylene glycol (PEG) radioimmunoprecipitation assay was developed for the detection of antibody to Haemophilus influenzae b capsular polysaccharide, polyribosylribitol phosphate (PRP). The radioactive antigen, [3H]PRP, with a high specific activity, was produced by growing the organism in the presence of [3H]ribose and was purified by hydroxylapatite and Sepharose 4B column chromatography. In the assay, PEG (12.5%) was used to separate antibody-bound [3H]PRP from free [3H]PRP. The assay covered the range of 0.5 and 20 ng antibody/assay at a maximum sensitivity of 0.5 approximately 1.0 ng antibody/assay. With various dilutions (1-20 ng antibody/assay) of S. Klein reference antiserum, the within-run coefficient of variation (CV) of 10 replicates ranged from 3.5 to 8.5%. Average CVs of 8.9% and 11.0% were obtained in the between-run and day-to-day reproducibility studies. The binding of [3H]PRP to S. Klein reference antiserum was severely inhibited by a minute amount of non-radioactive PRP; however, no significant interference was found in the presence of high concentrations of polysaccharides from Escherichia coli K100 and Streptococcus pneumoniae indicating that the RIA was highly specific for antibody to H. influenzae b PRP. The present RIA is a simple, specific, sensitive and reproducible procedure for the evaluation of antibody responses of young animals and infants to H. influenzae b vaccines and infections.
一种新的聚乙二醇（PEG）放射免疫沉淀测定法被开发出来，用于检测针对b型流感嗜血杆菌荚膜多糖——多聚核糖基核糖醇磷酸酯（PRP）的抗体。通过在[³H]核糖存在的情况下培养该生物体，产生了具有高比活性的放射性抗原[³H]PRP，并通过羟基磷灰石和琼脂糖4B柱色谱法进行纯化。在该测定法中，使用PEG（12.5%）将与抗体结合的[³H]PRP与游离的[³H]PRP分离。该测定法涵盖的抗体范围为0.5至20 ng/测定，最大灵敏度约为0.5至1.0 ng抗体/测定。对于S. Klein参考抗血清的各种稀释度（1 - 20 ng抗体/测定），10次重复测定的批内变异系数（CV）范围为3.5%至8.5%。在批间和日常重复性研究中，平均变异系数分别为8.9%和11.0%。微量的非放射性PRP会严重抑制[³H]PRP与S. Klein参考抗血清的结合；然而，在存在高浓度的大肠杆菌K100和肺炎链球菌多糖的情况下未发现明显干扰，这表明该放射免疫测定法对b型流感嗜血杆菌PRP抗体具有高度特异性。目前的放射免疫测定法是一种简单、特异、灵敏且可重复的方法，用于评估幼龄动物和婴儿对b型流感嗜血杆菌疫苗和感染的抗体反应。
