Dasch G A, Halle S, Bourgeois A L
J Clin Microbiol. 1979 Jan;9(1):38-48. doi: 10.1128/jcm.9.1.38-48.1979.
A microtiter enzyme-linked immunosorbent assay (ELISA) has been developed for the titration of antibodies against scrub typhus in human and animal sera. Scrub typhus rickettsiae were grown in monolayers of irradiated mouse LM3 cells and separated from host cell materials by differential centrifugation, filtration through a glass filter (AP-20, Millipore Corp.), and isopycnic banding in Renografin density gradients. The scrub typhus ELISA antigens were obtained from the purified viable rickettsiae by French pressure cell disruption and addition of 0.2% Formalin to the soluble extract. Antisera prepared in rabbits against the prototype Karp, the Kato, and the Gilliam strains of scrub typhus were used to standardize the ELISA and to compare its sensitivity and specificity to that of the indirect fluorescent antibody test (IFA). ELISA titers were measured as the greatest serum dilution showing an optical density 0.25 above controls or by the optical density achieved at a fixed serum dilution. The IFA and ELISA end point titers were quite similar, and all three measures of titer had comparable specificity for the strains of scrub typhus. No cross-reactions between the typhus and scrub typhus wera were observed by ELISA. Both the immunoglobulin M (IgM) and IgG antibody titers of 12 sequential sera from four patients with scrub typhus were obtained by IFA and ELISA. The IFA and ELISA end point titers for IgM and IgG had correlation coefficients of 0.91 and 0.97, respectively, whereas the ELISA optical density values at a serum dilution of 1:100 had slightly lower correlations with IFA titers (0.80 and 0.94). Early rising IgM titers followed by rising IgG titers were demonstrated by ELISA in three patients with primary scrub typhus infections, whereas the IgG response predominated in a patient with a reinfection. It is concluded that the ELISA for scrub typhus is a very satisfactory alternative to the IFA test.
已开发出一种微量滴定酶联免疫吸附测定法（ELISA），用于滴定人和动物血清中抗恙虫病的抗体。恙虫病立克次体在经辐照的小鼠LM3细胞单层中生长，通过差速离心、经玻璃滤器（AP - 20，密理博公司）过滤以及在泛影葡胺密度梯度中进行等密度离心，将其与宿主细胞物质分离。恙虫病ELISA抗原是通过法国压力细胞破碎法从纯化的活立克次体中获得的，并向可溶性提取物中加入0.2%的福尔马林。用兔制备的抗恙虫病原型株卡尔普、加藤和吉列姆株的抗血清对ELISA进行标准化，并将其敏感性和特异性与间接荧光抗体试验（IFA）进行比较。ELISA滴度的测量方法是：显示光密度比对照高0.25的最大血清稀释度，或在固定血清稀释度下达到的光密度。IFA和ELISA终点滴度非常相似，并且所有三种滴度测量方法对恙虫病株都具有相当的特异性。ELISA未观察到斑疹伤寒和恙虫病之间的交叉反应。通过IFA和ELISA获得了4例恙虫病患者12份连续血清的免疫球蛋白M（IgM）和IgG抗体滴度。IgM和IgG的IFA和ELISA终点滴度的相关系数分别为0.91和0.97，而在血清稀释度为1:100时ELISA的光密度值与IFA滴度的相关性略低（0.80和0.94）。ELISA在3例原发性恙虫病感染患者中显示出早期IgM滴度升高，随后IgG滴度升高，而在1例再感染患者中IgG反应占主导。结论是，恙虫病ELISA是IFA试验非常令人满意的替代方法。
