HIV-1 infection requires lifelong therapy with antiretroviral drugs due to the existence of a latent reservoir of transcriptionally inactive integrated proviruses. The goal of HIV-1 cure research is to eliminate or functionally silence this reservoir. To this end, there are numerous ongoing studies to evaluate immunological approaches, including monoclonal antibody therapies. Evaluating the results of these studies requires sensitive and specific measures of the reservoir. Here, we describe a relatively high-throughput combined quantitative PCR (qPCR) and next-generation sequencing method. Four different qPCR probes covering the packaging signal (PS), group-specific antigen (gag), polymerase (pol), and envelope (env) are combined in a single multiplex reaction to detect the HIV-1 genome in limiting dilution samples followed by sequence verification of individual reactions that are positive for combinations of any two of the four probes (Q4PCR). This sensitive and specific approach allows for an unbiased characterization of the HIV-1 latent reservoir.

由于存在转录失活的整合原病毒的潜在储存库，HIV-1 感染需要终生使用抗逆转录病毒药物进行治疗。 HIV-1 治疗研究的目标是消除或在功能上沉默该病毒库。为此，有许多正在进行的研究来评估免疫学方法，包括单克隆抗体疗法。评估这些研究的结果需要对储层进行敏感且具体的测量。在这里，我们描述了一种相对高通量的组合定量 PCR (qPCR) 和新一代测序方法。将涵盖包装信号 (PS)、组特异性抗原 ( gag )、聚合酶 ( pol ) 和包膜 ( env ) 的四种不同 qPCR 探针组合在一个多重反应中，以检测有限稀释样品中的 HIV-1 基因组，然后对四个探针中任意两个的组合呈阳性的单个反应进行序列验证 (Q4PCR)。这种敏感且特异的方法可以对 HIV-1 潜伏病毒库进行公正的表征。