Alexandre Kabamba, Malatji Kanyane, Mulaudzi Takalani
Council for Scientific and Industrial Research, Emerging Research Area Platform, Next Generation Health Cluster, Pretoria Gauteng, South Africa.
University of the Witwatersrand, School of Pathology, Johannesburg, South Africa.
MethodsX. 2020 Jul 26;7:101014. doi: 10.1016/j.mex.2020.101014. eCollection 2020.
HIV-1 infection is caused by cell-free and cell-associated viruses. Currently most of the assays used to screen potential HIV-1 entry inhibitors focus on the inhibition of cell-free viruses. One assay that is widely employed is the TZM-bl neutralization assay that uses pseudotyped viruses. However, a study by Abela et al. showed that many inhibitors that potently inhibit cell-free HIV-1 in this assay can be less effective against the cell-to-cell transmission of the virus. These researchers then designed a method to screen entry inhibitors for activity against cell-associated HIV-1, using pseudotyped viruses. The main limitation of this method, however, was that it can only be reliably employed against viruses that cannot infect target cells as cell-free virion in the absence of a polycation supplement such as DEAE (diethylaminoethyl). Thus, in the current study we provide modifications to this method that solves the problem and makes it possible to study entry inhibitors against cell-to-cell infection of both polycation depend and independent viruses. The main modification involves the introduction of the relative light unit (RLU) vs. virus producing 293-T cells / corresponding supernatants graph. This graph is used to select a virus input that only allows for the detection of cell-associated viruses infection.•The method is a modification of the cell-to-cell transmission assay published by Abela et al.•The method allows for the study of the inhibition of cell-to-cell transmission of both polycation dependent and independent HIV-1 pseudoviruses.
HIV-1感染由游离病毒和细胞相关病毒引起。目前，大多数用于筛选潜在HIV-1进入抑制剂的检测方法都集中在对游离病毒的抑制上。一种广泛使用的检测方法是使用假型病毒的TZM-bl中和检测法。然而，阿贝拉等人的一项研究表明，许多在该检测中能有效抑制游离HIV-1的抑制剂对病毒的细胞间传播可能效果较差。这些研究人员随后设计了一种方法，使用假型病毒筛选针对细胞相关HIV-1具有活性的进入抑制剂。然而，该方法的主要局限性在于，它只能可靠地用于在没有诸如DEAE（二乙氨基乙基）等聚阳离子补充剂的情况下不能作为游离病毒粒子感染靶细胞的病毒。因此，在本研究中，我们对该方法进行了改进，解决了这个问题，并使得研究针对聚阳离子依赖型和非依赖型病毒细胞间感染的进入抑制剂成为可能。主要改进包括引入相对光单位（RLU）与病毒产生细胞293-T/相应上清液的图表。该图表用于选择仅允许检测细胞相关病毒感染的病毒输入量。
•该方法是对阿贝拉等人发表的细胞间传播检测法的改进。
•该方法可用于研究聚阳离子依赖型和非依赖型HIV-1假病毒细胞间传播的抑制作用。
