Farzadegan H, Imagawa D, Gupta P, Lee M H, Jacobson L, Saah A, Grovit K, Rinaldo C R, Polk B F
Johns Hopkins University School of Hygiene and Public Health, Baltimore, Maryland 21205.
J Acquir Immune Defic Syndr (1988). 1990;3(10):981-6.
A multicenter study was undertaken to determine the sensitivity and reproducibility of markers for human immunodeficiency virus type 1 (HIV-1) viral growth and the effect of various preparations of lymphocytes on the sensitivity of standard and routinely used procedures for HIV-1 isolation. In phase 1, cocultivated culture supernatants obtained from 10 HIV-1 cultures were transported to three Multicenter AIDS Cohort Study (MACS) Virology Laboratories. Three commercial HIV-p24 antigen capture (AC) tests and two reverse transcriptase (RT) assays were used to ascertain the replication of HIV-1. The Du Pont and Abbott AC assays were found to be most sensitive (85-100%), and the RT assay with 24-h incubation period had comparable sensitivity (75-100%). In phase II, the sensitivity of standard cocultivation procedure for HIV-1 isolation was compared using freshly phytohemagglutinin-P (PHA-P)-stimulated, stimulated-frozen, and frozen-thawed and then stimulated normal human peripheral blood mononuclear cells (PBMCs) as cocultivating cells. Blood samples from 13 HIV-1 infected individuals with various CD4+ cell counts were cocultivated in each of the three MACS laboratories using one of the aforementioned normal PBMCs. The PHA-P-stimulated fresh normal PBMC showed a maximum isolation rate of 100% (13 of 13) with an average of 8 days to positivity. This rate of isolation was significantly greater than other rates using any one of the other PBMC preparations. These findings demonstrated that the use of freshly PHA-P stimulated PBMCs maximized HIV-1 isolation from blood when a sensitive HIV-1 p24 AC assay or RT assay with overnight incubation is employed for the detection of HIV in culture supernatant.
开展了一项多中心研究，以确定1型人类免疫缺陷病毒（HIV-1）病毒生长标志物的敏感性和可重复性，以及各种淋巴细胞制剂对HIV-1分离标准和常规使用程序敏感性的影响。在第1阶段，从10个HIV-1培养物中获得的共培养上清液被运送到三个多中心艾滋病队列研究（MACS）病毒学实验室。使用三种商业HIV-p24抗原捕获（AC）试验和两种逆转录酶（RT）测定法来确定HIV-1的复制情况。发现杜邦和雅培AC试验最敏感（85%-100%），孵育24小时的RT测定法具有相当的敏感性（75%-100%）。在第II阶段，使用新鲜的植物血凝素-P（PHA-P）刺激、刺激后冷冻、冷冻解冻后再刺激的正常人外周血单核细胞（PBMC）作为共培养细胞，比较了HIV-1分离标准共培养程序的敏感性。来自13名不同CD4+细胞计数的HIV-1感染个体的血样在三个MACS实验室中的每一个中，使用上述正常PBMC之一进行共培养。PHA-P刺激的新鲜正常PBMC显示出最高的分离率为100%（13例中的13例），平均阳性时间为8天。该分离率显著高于使用其他任何一种PBMC制剂的分离率。这些发现表明，当使用敏感的HIV-1 p24 AC试验或过夜孵育的RT试验检测培养上清液中的HIV时，使用新鲜PHA-P刺激的PBMC可使从血液中分离HIV-1的效率最大化。
