(C) Schematic of the Nluc and GFP co-expressing retroviral vector pMIG.Nluc.IRES.GFP. (p24+) population 18 hours and 24 hours post-infection in CEMSS.NKr.R5 cells infected with (D) BG505.GFP or (E) BG505.GFP* T/F reporter viruses.(TIF) ppat.1008161.s001.tif (4.0M) GUID:?E8AB5A50-9A55-47B0-A585-4BBE40145F6E S2 Fig: In vivo reporter gene stability in Hu-PBL mice. (A-C) GFP reporter gene stability in p24+ CD3+ PBMCs extracted from the peripheral blood of Hu-PBL mice infected intraperitonially (i.p.) with 1 x 107 infectious units (IUs) HIV-1 T/F reporter virus. (A) Representative FACS plots gated on CD3+ PBMCs extracted from BG505.GFP* infected Hu-PBL mice (n = 4). Data is representative of four individual Hu-PBL mice. (B-C) Average GFP reporter gene stability in Hu-PBL mice infected with 1 x 107 infectious units (IUs) of BG505.GFP* (n = 4) (B), and BG505.GFP (n = 3) (C) T/F reporter virus for 14C16 days. Data displayed as the percentage of GFP and p24 double-positive cells in the total p24+ population. A line crosses the average percent GFP expressing cells within the total p24+ cell population for mice analyzed at each time point.(TIF) ppat.1008161.s002.tif (1.7M) GUID:?53319162-A070-4912-8C35-94961BC32D2E S3 Fig: Cryoimmunofluroescent and LS-MPM intravital spleen imaging of Hu-PBL mice injected i.p. with 1 x 107 IUs TRJO.GFP 7 days post-infection. (A) Cryoimmunofluorescent confocal imaging of splenic tissue sections; areas with GFP expressing cells are magnified in panels 1 and 2. White arrows indicate putative syncytia formed during infection. (B-G) LS-MPM imaging of Gadoxetate Disodium spleen tissue from a Hu-PBL mouse injected i.p. with 1 x 107 IUs TRJO.GFP 7 days post-infection and injected with RFP expressing CD4 T cells 24 hours prior to imaging. (B,C) LS-MPM intravital imaging of an area in the spleen with GFP expressing cells. A representative cell exhibiting long membrane extensions is outlined in white dashes (B) with motion tracks of GFP expressing cells in (C). (D-E) LS-MPM image of GFP and CD4 co-expressing syncytium in the spleen of a TRJO.GFP-infected Hu-PBL mouse (D) and the same image with CD4 expression alone (E). (F-G) LS-MPM image of GFP expressing cells in the spleen as in (D) with a GFP and CD4 co-expressing Gadoxetate Disodium cell indicated by the white arrow and CD4 expressing cells alone (G). All scale bars correspond to 100 m.(TIF) ppat.1008161.s003.tif (2.9M) GUID:?4C8B2F1C-70C6-4948-AE78-A29AB08E3EFB S4 Fig: RNA viral load assay and SG-PERT RT activity assay sensitivities. (A) Peripheral blood mononuclear cell (PBMC) derived HIV-1 JR-CSF viral supernatant was stored in independent aliquots of equivalent volume in order to compare the sensitivity of the Quantitect qRT-PCR viral weight assay and the SG-PERT reverse transcriptase activity qPCR assay in parallel. (B) The Quantitect qRT-PCR viral weight assay and the SG-PERT reverse transcriptase activity qPCR assay was run in parallel with viral RNA eluate and HIV-1 supernatant serially diluted until the limit of detection for each assay was reached. Data demonstrated as the average cycle threshold (Cq) ideals identified from two technical replicates at each dilution. The limit of detection was defined as the Cq value at which the linear range of the assay ended. Complete quantification of HIV-1 particles was identified from a viral RNA standard curve run in parallel with the Quantitect qRT-PCR viral weight assay.(TIF) ppat.1008161.s004.tif (940K) GUID:?963745E8-9C13-49BC-9801-250369E2C15C S5 Fig: Longitudinal NNT1 non-invasive bioluminescent imaging of HIV-1 acute infection, suppression, and recrudescent infection in the Hu-BLT mouse group placed on cART 12 days post-infection. (A) Bioluminescent imaging of distributing illness of Hu-BLT Mouse #3 infected with 1 x 106 Gadoxetate Disodium IUs of Q23.BG505.Nluc T/F reporter disease and placed on a daily cART routine comprised of daily i.p. cART injections of Truvada and Isentress 12 days post-infection. (B) Plasma reverse transcriptase activity from Hu-BLT Mouse #3 (above) and whole-animal Nluc transmission (below) over the course of the 40-day time imaging period. Plasma reverse transcriptase activity in serum samples taken every six days over the course of the imaging period was measured via the SG-PERT reverse transcriptase activity assay and described as reverse transcriptase activity devices / mL above endogenous uninfected background levels (dotted collection). (C) Whole animal ex vivo necroscopic analysis of rebounding illness in Hu-BLT.