Haimovici F, Mayer K H, Anderson D J
Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
J Acquir Immune Defic Syndr Hum Retrovirol. 1997 Jul 1;15(3):185-91. doi: 10.1097/00042560-199707010-00001.
A quantitative enzyme-linked immunosorbent assay (ELISA) has been developed for the quantitation of HIV-1-specific immunoglobulins of the IgG, IgA, and IgM isotypes and was used to assess HIV-specific antibody concentrations in semen and cervicovaginal lavage (CVL) specimens. Immunoglobulin isotype concentrations were assessed by capture ELISA using immunoglobulin isotype-specific secondary antibodies and commercial IgG, IgA, and IgM standards. HIV-1 antibody detection kits (Abbott Laboratories, North Chicago, IL, U.S.A.) and immunoglobulin isotype-specific secondary antibodies were used to obtain optical density (OD) units for HIV-1-specific antibodies of each isotype. To determine the antibody concentrations from the OD values, ODs were compared with those from HIV-1-specific antibody isotype standards of known concentration obtained from CVL specimens of seropositive women by affinity binding to HIV antigen-coated beads and acid elution. The sensitivity of the HIV-1-specific immunoglobulin assay was 0.01 microg/ml for IgG, 0.04 microg/ml for IgA, and 0.08 microg/ml for IgM. The interassay coefficient of variation for the different immunoglobulin isotypes varied from 5% to 33%, being lowest for IgG and highest for IgM. HIV-1-specific antibodies were detected in all semen samples from seropositive men in concentrations ranging from 53 to 261 microg/ml. Thirteen of 14 samples contained high levels of HIV-1-specific IgG antibodies (22-72 microg/ml) whereas 10 of the 14 (71%) semen samples contained detectable but lower levels of HIV-1-specific IgA and IgM (maximum level: 3.7 microg/ml for IgA and 14.8 microg/ml for IgM). HIV-1-specific IgG antibodies were detected in all 196 CVL samples from seropositive women in concentrations ranging from 0.01 to 47 microg/ml, whereas only 16 women (8%) had IgA levels above the detectable limit (range, 1.4-3.9 microg/ml), and only eight women (4%) had IgM levels above the detectable limit (range, 1.8-8.6 microg/ml). None of 80 low-risk women or 20 low-risk men (negative controls) had detectable levels of HIV-1-specific antibodies in genital tract specimens. HIV-1-specific IgG levels in CVL specimens of seropositive women were significantly higher in individuals who had acquired HIV through heterosexual transmission, and a majority of women with elevated levels of HIV-specific IgA isotype antibodies in CVL samples had evidence of genital tract inflammation (>10[4] polymorphonuclear leukocytes [PMNs]/ml). Use of this quantitative method will facilitate direct comparison of data obtained within and among laboratories and enable further research on factors affecting antibody levels in genital tract secretions and their effects on HIV-1 transmission.
已开发出一种定量酶联免疫吸附测定法（ELISA），用于定量检测HIV-1特异性IgG、IgA和IgM同种型免疫球蛋白，并用于评估精液和宫颈阴道灌洗液（CVL）标本中HIV特异性抗体浓度。通过使用免疫球蛋白同种型特异性二抗和商业化IgG、IgA及IgM标准品的捕获ELISA来评估免疫球蛋白同种型浓度。使用HIV-1抗体检测试剂盒（美国伊利诺伊州北芝加哥雅培实验室）和免疫球蛋白同种型特异性二抗来获取每种同种型HIV-1特异性抗体的光密度（OD）值。为了根据OD值确定抗体浓度，将OD值与通过与HIV抗原包被的珠子亲和结合及酸洗脱从血清反应阳性女性的CVL标本中获得的已知浓度的HIV-1特异性抗体同种型标准品的OD值进行比较。HIV-1特异性免疫球蛋白测定法对IgG的灵敏度为0.01μg/ml，对IgA为0.04μg/ml，对IgM为0.08μg/ml。不同免疫球蛋白同种型的测定间变异系数在5%至33%之间，其中IgG最低，IgM最高。在血清反应阳性男性的所有精液样本中均检测到HIV-1特异性抗体，浓度范围为53至261μg/ml。14个样本中有13个含有高水平的HIV-1特异性IgG抗体（22 - 72μg/ml），而14个精液样本中有10个（71%）含有可检测到但水平较低的HIV-1特异性IgA和IgM（IgA最高水平为3.7μg/ml，IgM为14.8μg/ml）。在血清反应阳性女性的所有196个CVL样本中均检测到HIV-1特异性IgG抗体，浓度范围为0.01至47μg/ml，而只有16名女性（8%）的IgA水平高于可检测限（范围为1.4 - 3.9μg/ml），只有8名女性（4%）的IgM水平高于可检测限（范围为1.8 - 8.6μg/ml）。80名低风险女性或20名低风险男性（阴性对照）的生殖道标本中均未检测到可检测水平的HIV-1特异性抗体。通过异性传播感染HIV的个体，其血清反应阳性女性CVL标本中的HIV-1特异性IgG水平显著更高，并且CVL样本中HIV特异性IgA同种型抗体水平升高的大多数女性有生殖道炎症的证据（>10[4]个多形核白细胞[PMN]/ml）。使用这种定量方法将有助于直接比较不同实验室内部和之间获得的数据，并能够进一步研究影响生殖道分泌物中抗体水平的因素及其对HIV-1传播的影响。
