Rucheton M, Graafland H, Fanton H, Ursule L, Ferrier P, Larsen C J
Virology. 1985 Jul 30;144(2):468-80. doi: 10.1016/0042-6822(85)90287-9.
An immunoblotting procedure using viral proteins from purified murine sarcoma virus or MSV-(MLV) has been developed to characterize antiviral antibodies in sera from patients with autoimmune connective tissue disorders. Fifty-eight sera with anti-Sm, anti-RNP, anti-SS-B (La), and other undefined specificities were found to react with several major viral polypeptide bands. Most of them corresponded to gag-gene-encoded products: pr65gag, p40gag, p30, p15, p12 and p10. Other bands with molecular weights averaging 90K, 60K, 45K, and 28K were recognized by a few sera. Immunological specificity of the reaction was assessed by reproducing the tests with IgG purified from sera and from corresponding F(ab')2 fragments. Moreover, the specificity of the reaction with gag proteins was confirmed by repeating the tests with p30 and p15 prior purified by immunoprecipitation with anti-p30 and anti-p15 goat sera. Furthermore, the gag polypeptides were recognized by human sera by replacing MSV-(MLV) by three other murine retroviruses of different origin. An indirect confirmation of these results was obtained by applying this method to sera of MRL lpr/lpr mice which develop an autoimmune syndrome comparable to that of human systemic lupus erythematosus. In agreement with previously published results (C. Rordorf, C. Gambke, and J. Gordon (1983), J. Immunol. Methods 59, 105-112), we found that anti-gag-gene antibodies were present in the sera of individual mice. Patterns of reactivity were found to vary with the age of the animals. No retroviral polypeptide was significantly detected in the great majority (80%) of sera from normal donors. However, 5 out of 25 sera showed faint bands although to a lesser extent than pathological sera. These five sera also reacted with HeLa cell purified HnRNPs, suggesting that their normal status should be reconsidered.
已开发出一种免疫印迹程序,该程序使用来自纯化的鼠肉瘤病毒或MSV-(MLV)的病毒蛋白,以鉴定自身免疫性结缔组织疾病患者血清中的抗病毒抗体。发现58份具有抗Sm、抗RNP、抗SS-B(La)及其他未明确特异性的血清与几条主要的病毒多肽带发生反应。其中大多数对应于gag基因编码产物:pr65gag、p40gag、p30、p15、p12和p10。少数血清识别出其他分子量平均为90K、60K、45K和28K的条带。通过用从血清和相应F(ab')2片段纯化的IgG重复试验来评估反应的免疫特异性。此外,通过用抗p30和抗p15山羊血清免疫沉淀预先纯化的p30和p15重复试验,证实了与gag蛋白反应的特异性。此外,通过用另外三种不同来源的鼠逆转录病毒替代MSV-(MLV),人血清可识别gag多肽。通过将该方法应用于发生与人类系统性红斑狼疮相当的自身免疫综合征的MRL lpr/lpr小鼠血清,间接证实了这些结果。与先前发表的结果一致(C. Rordorf、C. Gambke和J. Gordon(1983年),《免疫学方法杂志》59,105 - 112),我们发现个别小鼠血清中存在抗gag基因抗体。发现反应模式随动物年龄而变化。在绝大多数(80%)正常供体血清中未明显检测到逆转录病毒多肽。然而,25份血清中有5份显示出微弱条带,尽管程度低于病理血清。这五份血清也与HeLa细胞纯化的hnRNP发生反应,表明应重新考虑它们的正常状态。