Calculating HIV-1 Infectious Titre Using a Virtual TCID50 Method

Studies of HIV-1 replication kinetics and fitness require an accurate determination of the level of infectious HIV-1 present in virus stocks. The standard technique for measuring the level of replication-competent infectious virus in culture supernatants or patient samples is the tissue culture dose for 50% infectivity (TCID₅₀ ), which provides an accurate assessment of the level of infectious HIV-1. However, it is a time-consuming technique which typically takes two or more weeks to complete and requires PHA-stimulated PBMC from HIV-1 seronegative donors or an appropriate cell line. Thus rapid, cell-free surrogate measures for TCID₅₀ are desirable. Here, we introduce the virtual TCID₅₀ technique: a new cell-free method estimating a surrogate of infectious titer by comparing the reverse transcriptase activity in virus stock to that of reference viruses with a known TCID₅₀ value. We have demonstrated that the virtual TCID₅₀ obtained through this technique is comparable to the actual infectious TCID₅₀ . This method greatly simplifies the process of accurate HIV-1 titration and is particularly beneficial for studies which require titration of large number of HIV-1 isolates.