Heterologous Prime-Boost HIV-1 Vaccination Regimens in Pre-Clinical and Clinical Trials - PubMed (original) (raw)

Figure 1.

Durable antibody responses elicited by successive HIV-1 envelope immunizations with three different recombinant delivery systems. Female C57BL/6 mice were purchased from the Jackson Laboratory (Bar Harbor, ME) and housed under specific pathogen-free conditions in a BL1 or BL2+ containment area at St. Jude Children’s Research Hospital animal facilities, with adherence to AAALAC Guidelines. Mice were grouped (n = 3) for immunizations with UG92005 (UG) envelope using recombinant vectors: (i) DNA (‘D’, 100 μg, carrying a gp140 envelope sequence) administered i.m. in the gastrocnemius muscles, (ii) CHO-derived recombinant UG envelope protein (‘P’, 2 μg, 100 μl gp140 envelope in PBS mixed with 100 μl CFA) administered i.p., and (iii) SeV (‘S’, 104 plaque forming units, carrying a gp120 envelope sequence for expression in infected cells) administered i.n.. Groups of mice received one, two or three of the recombinant vectors (or no vaccine, ‘0’), administered with one month intervals. Eight months after the completion of inoculations, ELISAs were performed with serially diluted serum samples. Antibody binding titers were determined using Prism Software (Non-Linear Regression, GraphPad Prism®, San Diego, CA). Mean titers and standard errors are shown. Some data points represent fewer than 3 animals due to mouse death or non-conformance of data to the non-linear regression curve (O-S-D, O-D-P, O-P-D, O-S-P, S-P-D, P-S-D). To perform the ELISA, 96-well plates (BD Biosciences, Franklin Lakes, NJ) were coated overnight at 4 °C with 2 μg/ml of purified CHO-derived UG gp140 envelope protein in PBS. The plates were washed three times with 0.05% Tween 20 in PBS, blocked with 1% BSA/PBS at room temperature for 1 h, and washed an additional three times. Samples were serially diluted in PBS to a final volume of 50 μl and were incubated in wells for 2 h at room temperature. After three washes, alkaline phosphatase-conjugated anti-mouse IgG1 (50 μl/well; Southern Biotechnology Associates, Birmingham, AL) diluted 1/1000 in 1% BSA/0.05% Tween 20/PBS was added for 1 h at room temperature. Following five washes, the assay was developed with 75 μl/well of p-nitrophenyl phosphate (Sigma-Aldrich) substrate (1 mg/ml in diethanolamine buffer) and was read at OD450nm after 1 hour at 37 °C.