Department of Pathology, University of Miamigrid.26790.3a Miller School of Medicine, Miami, Florida, USA.
Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, Wisconsin, USA.
J Virol. 2022 Jan 26;96(2):e0158221. doi: 10.1128/JVI.01582-21. Epub 2021 Nov 3.
BG505 SOSIP.664 (hereafter referred to as SOSIP), a stabilized trimeric mimic of the HIV-1 envelope spike resembling the native viral spike, is a useful tool for isolating anti-HIV-1 neutralizing antibodies. We screened long-term SHIV-AD8 infected rhesus monkeys for potency and breadth of serum neutralizing activity against autologous and heterologous viruses: SHIV-AD8, HIV-1 YU2, HIV-1 JR-CSF, and HIV-1 NL4-3. Monkey rh2436 neutralized all viruses tested and showed strong reactivity to the SOSIP trimer, suggesting this was a promising candidate for attempts at monoclonal antibody (MAb) isolation. MAbs were isolated by performing single B-cell sorts from peripheral blood mononuclear cells (PBMC) by FACS using the SOSIP trimer as a probe. An initial round of sorted cells revealed the majority of isolated MAbs were directed to the gp41 external domain portion of the SOSIP trimer and were mostly non-neutralizing against tested isolates. A second sort was performed, introducing a gp41 blocking step prior to PBMC staining and FACS sorting. These isolated MAbs bound SOSIP trimer but were no longer directed to the gp41 external domain portion. A significantly higher proportion of MAbs with neutralizing activity were obtained with this strategy. Our data show this pre-blocking step with gp41 greatly increases the yield of non-gp41-reactive, SOSIP-specific MAbs and increases the likelihood of isolating MAbs with neutralizing activity. Recent advancements in the field have focused on the isolation and use of broadly neutralizing antibodies for both prophylaxis and therapy. Finding a useful probe to isolate broad potent neutralizing antibodies while avoiding non-neutralizing antibodies is important. The SOSIP trimer has been shown to be a great tool for this purpose because it binds known broadly neutralizing antibodies. However, the SOSIP trimer can isolate non-neutralizing antibodies as well, including gp41-specific MAbs. Introducing a pre-blocking step with gp41 recombinant protein decreased the percent of gp41-specific antibodies isolated with SOSIP probe, as well as increased the number of neutralizing antibodies isolated. This method can be used as a tool to increase the chances of isolating neutralizing antibodies.
BG505 SOSIP.664（以下简称 SOSIP）是一种稳定的 HIV-1 包膜刺突三聚体模拟物，类似于天然病毒刺突，是分离抗 HIV-1 中和抗体的有用工具。我们筛选了长期感染 SHIV-AD8 的恒河猴的血清中和活性的效力和广度，以针对同源和异源病毒：SHIV-AD8、HIV-1 YU2、HIV-1 JR-CSF 和 HIV-1 NL4-3。猴 rh2436 中和了所有测试的病毒，并对 SOSIP 三聚体表现出强烈的反应性，这表明它是分离单克隆抗体（MAb）的有希望的候选者。通过使用 SOSIP 三聚体作为探针，通过 FACS 对外周血单核细胞（PBMC）进行单个 B 细胞分选，分离 MAb。第一轮分选细胞揭示了大多数分离的 MAb 针对 SOSIP 三聚体的 gp41 外部结构域部分，并且大多数对测试分离物没有中和作用。进行了第二轮分选，在 PBMC 染色和 FACS 分选之前引入 gp41 阻断步骤。这些分离的 MAb 结合 SOSIP 三聚体，但不再针对 gp41 外部结构域部分。使用这种策略获得了具有中和活性的 MAb 的比例显著增加。我们的数据表明，gp41 的这种预阻断步骤大大增加了非 gp41 反应性、SOSIP 特异性 MAb 的产量，并增加了分离具有中和活性的 MAb 的可能性。该领域的最新进展集中在分离和使用广谱中和抗体用于预防和治疗。找到一种有用的探针来分离广谱有效中和抗体，同时避免非中和抗体是很重要的。SOSIP 三聚体已被证明是一种很好的工具，因为它可以结合已知的广谱中和抗体。然而，SOSIP 三聚体也可以分离非中和抗体，包括 gp41 特异性 MAb。用 gp41 重组蛋白引入预阻断步骤降低了用 SOSIP 探针分离的 gp41 特异性抗体的百分比，同时增加了分离的中和抗体的数量。该方法可用作增加分离中和抗体的机会的工具。
