Lentiviral technology has proven to be a powerful tool to express exogenous genes in dividing and non-dividing cells. Currently, most protocols for generating high-titer lentivirus require ultracentrifugation, which can be an instrumental barrier for routine operations in a laboratory. In this study, the effect of relative centrifugal force (RCF) on the concentration efficiency of the lentivirus was systematically explored, and it was found that sucrose gradient centrifugation with a relatively low speed (≤10,000 g) robustly produces a high-titer virus (up to 2 × 108 TU/ml). The optimal sucrose concentration is 10%, and the recovery rate of the functional virus is greater than 80%. The infection efficiency of both concentrated and un-concentrated lentivirus decreases rapidly when the viruses are stored at 4 °C (τ ≈ 1.3 days) or subjected to multiple freeze-thaw cycles (τ = 1.1 rounds). In summary, we describe an efficient and easy-to-handle protocol for high-titer lentivirus purification.
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机译：慢病毒技术已被证明是在分裂和非分裂细胞中表达外源基因的强大工具。当前，大多数产生高滴度慢病毒的方案都需要超速离心，这可能是实验室常规操作的工具性障碍。在这项研究中，系统地研究了相对离心力（RCF）对慢病毒的浓缩效率的影响，并且发现以相对较低的速度（≤10,000g）进行蔗糖梯度离心可以稳固地产生高滴度病毒（最高2×10 8 TU / ml）。最佳蔗糖浓度为10％，功能病毒的回收率大于80％。当病毒在4°C（τ≈1.3天）或经受多个冻融循环（τ= 1.1轮）储存时，浓缩和未浓缩慢病毒的感染效率都会迅速降低。总之，我们描述了一种高效且易于处理的高滴度慢病毒纯化方案。
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