Innotest hiv 1 antigen mab

The freshly sorted Treg cells were activated with Dynabeads Human Treg Expander (Invitrogen) following manufacturer’s recommendations and treated for 24 hours with dendrimers. The treated and non-treated Treg cells were then infected with X4-tropic HIV-1_NL4.3 (MOI of 0.1) for 3 hours. These cells were then extensively washed and cultured in the presence of 500 U/ml of rIL-2. The controls of inhibition of HIV-1 replication were performed with zidovuline (AZT) (Retrovir; GlaxoSmithKline, Brentford, UK).
The HIV-1 entry and infection were confirmed by measuring the concentration of p24^gag in the culture supernatant by ELISA (Innotest HIV-1 antigen mAb; Innogenetics, Gent, Belgium) or by the intracellular expression of p24 using the KC57 antibody.

The inhibition of HIV-1 transfer from Langerhans cells to T cells was evaluated using monocyte-derived Langerhans cells (LCs) obtained from PBMCs, and autologous CD4⁺T cells as we previously described [12 (link)]. Briefly, LCs were incubated with either clade of HIV-1 for 2hrs at 37°C, washed extensively to remove the free virus and distributed in 96 well plates at 100,000 cells / well. Indicated concentrations of FabA or G were then added to the corresponding wells. Finally, either medium alone, or resting CD4⁺T-cells in medium (100,000 cells / well) were added. The LC-T-cell co-cultures were incubated at 37°C for 5 days. Irrelevant Fab-IgA and Fab-IgG at similar concentrations showing a value between 3–7% depending of the clade were used as negative controls. Virus transfer was evaluated by measuring Gag-p24 released in the culture medium using a commercial ELISA (Innotest HIV-1 Antigen mAb, Innogenetics) according to manufacturer instructions. Results are expressed as % inhibition of transfer using formula [(LC+T-cell-Ab)—(LC+T-cell+Ab) / (LC+T-Ab)] x100.

A stock of HIV-1_JR-CSF (clade B, R5 tropic) was prepared on a large scale by transfecting 293T cells with a plasmid containing the DNA sequence of JR-CSF (NIH, Germantown, MD USA) [3 (link)]. The cell culture supernatant was concentrated, separated into single-use aliquots and stored at –80°C.
The HIV-1 primary isolates 92UG031 (clade A, R5) and 92BR025 (clade C, R5), obtained through the NIH AIDS Reagent Program, was amplified on PBMCs, as previously described [3 (link)]. Virus concentration was quantified by measuring p24 antigen by ELISA (Innotest HIV-1 Antigen mAb, Innogenetics).