Background

Evaluations of HIV curative interventions require reliable and efficient quantification of replication-competent latent reservoirs (LR). The “classic” quantitative viral outgrowth assay (QVOA) has been regarded as “gold standard,” although prohibitively resource- and labor-intensive. We compared six “next-gen” VOA employing PCR or ultrasensitive p24 to assess their suitability as scalable proxies for QVOA.

Methods

Next-gen VOA were compared to classic QVOA using single leukapheresis-derived samples from five ART-suppressed HIV⁺ participants and one HIV^- control; each lab tested blinded batches of three frozen and one fresh sample. Markov chain Monte Carlo methods estimated extra-Poisson variation at aliquot, batch, and lab levels. Models also estimated the effect of testing frozen versus fresh samples.

Results

Next-gen VOA had similar estimates of variation to QVOA. Assays with ultrasensitive readout reported higher IUPM than classic QVOA. Within-batch testing had 2.5-fold extra-Poisson variation (95%CI 2.1,3.5) for next-gen assays. Between-lab variation increased extra-Poisson variation to 3.4-fold (95% CI 2.6,5.4). Frozen storage did not substantially alter IUPM (–18%(–52%,+39%)).

Conclusions

The data offer cautious support for use of next-gen VOA as proxies for more laborious QVOA, while providing greater sensitivities and dynamic ranges. Measurement of LR in eradication strategies would benefit from high throughput and scalable assays.

中文翻译：

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评估下一代病毒生长试验作为经典 QVOA 测量 HIV-1 潜伏库大小的代理的适用性。

背景

对 HIV 治愈性干预措施的评估需要对具有复制能力的潜在水库 (LR) 进行可靠和有效的量化。“经典”定量病毒生长试验 (QVOA) 已被视为“黄金标准”，尽管资源和劳动密集型令人望而却步。我们比较了六种使用 PCR 或超灵敏 p24 的“下一代”VOA，以评估它们作为 QVOA 可扩展代理的适用性。

方法

^{使用来自 5 名 ART 抑制的 HIV +}参与者和 1 名 HIV ^-对照的单一白细胞分离术衍生样本，将下一代 VOA 与经典 QVOA 进行了比较；每个实验室都测试了三个冷冻样品和一个新鲜样品的盲批次。马尔可夫链蒙特卡罗方法估计了等分试样、批次和实验室级别的额外泊松变异。模型还估计了测试冷冻样品与新鲜样品的效果。

结果

下一代 VOA 对变异的估计与 QVOA 相似。具有超灵敏读数的测定报告的 IUPM 高于经典 QVOA。对于下一代测定，批次内测试具有 2.5 倍的额外泊松变异 (95%CI 2.1,3.5)。实验室间变异将额外泊松变异增加到 3.4 倍 (95% CI 2.6,5.4)。冷冻储存并没有显着改变 IUPM (–18%(–52%,+39%))。

结论

这些数据为使用下一代 VOA 作为更费力的 QVOA 的代理提供了谨慎的支持，同时提供了更大的灵敏度和动态范围。根除策略中的 LR 测量将受益于高通量和可扩展的测定。