Journal: International Journal of Clinical and Experimental Pathology
Article Title: Tetraspanin CD63 is a regulator of HIV-1 replication
Figure Lengend Snippet: CD63 mRNA down regulation by siRNA in human MDMs which are challenged with different concentration of HIV-1. A. MDMs (5 × 10 5 cells/well) were plated in 24-well plates and transfected with 50 nM siRNAs (CD63, CD4, and ERBB2IP). For controls, cells were treated with AZT (1 mM) or raltegravir (20 mM). Controls also included untreated cells or cells infected with HIV-1 SX (m.o.i. = 0.02) only. Total mRNA was isolated from each well using the Qiagen RNeasy kit 48 h after transfection. Quantitative RT-PCR was used to determine relative CD63 expression levels, normalizing to GAPDH expression as an internal control. B. MDMs were challenged with different concentration of HIV-1 SX (m.o.i. = 0.6, 0.2. 0.06 and 0.02, respectively). Supernatants were harvested for p24 detection on day 7 post-infection for MDMs using p24 Capture ELISA kit (ImmunoDiagnostics, Woburn, MA).
Article Snippet: Total mRNA was isolated from siRNA-transfected cells and MDMs using RNeasy Mini Kits (Qiagen).
Techniques: Concentration Assay, Transfection, Infection, Isolation, Quantitative RT-PCR, Expressing, Enzyme-linked Immunosorbent Assay