Fig 1: Specific Effect Of Stably Expressed Tox4 And Nova1 On Hiv-1/Vsv-G But Not On Mlv/Vsv-G Infection.A) 293T cells transduced with LVs carrying GFP-TOX4 PIR or GFP-TOX4 HMG or GFP-NOVA1 PIR or GFP- IBD (LEDGF) were analysed by FACS two weeks after transduction. 50,000 cells transduced with LVs were infected with B) 20 ng of p24 of HIV-1-Luc and C) with MLV-Luc, analyzed by luciferase assay and normalyzed by total proteins. D) Expression of EGFP-fusion proteins in infected cells. 10 µg of total cell extracts used for the infections, were separated by SDS-8% PAGE, and the presence or EGFP tagged proteins was analysed by western blotting of total extracts using an anti-GFP antibody (Abcam ab290) and normalized by actin (Abcam A5441).
Fig 2: Enrichment of GFP-tagged proteins by AptA–MS. (A) Schematic representation of the experimental design. (B) Cellular lysates prepared from HCT116 cells transfected with GFP or HSF1-GFP expressing plasmids were analyzed by anti-GFP (green) and anti-Actin (red, loading control) western blot. GFP (Abcam, ab290) and Actin (Sigma, MAB1501) antibodies were used at 1:2000 and 1:5000 dilutions, respectively. (C) Lysate from cells expressing GFP or HSF1-GFP were precipitated with the GFP- or Control (Ctrl)-aptamer and eluates were analyzed by gel electrophoresis and silver-staining. Bottom panel shows a fluorescence image of the eluates. (D) Enrichment analysis of HSF1 in AptA–MS samples from cells expressing GFP or HSF1-GFP, before or after heat shock pulled-down with the GFP- or the control-aptamer. Plot represents data from five independent biological replicates.
Fig 3: Evidence of SST4 expression in SST4-eGFP knockin mice. SST4 was detected immunohistochemically in tissue sections from SST4-eGFP knockin mice using a commercial rabbit anti-eGFP antibody (ab290, Abcam, Cambridge, UK; 1:1000). Scale bar: (A–E) = 60 µm; (F) = 100 µm.
Fig 4: Microdystrophin delivery in vivo. A 1-year-old female mouse with DMD (mdx4Cv) was treated in both legs with 50 μL of concentrated polyplex (12.5 µg pµDys in the left leg, 6.25 µg pµDys in the right leg). After 48 h, protein analysis was performed on leg tissues. (a) Western blot analysis of the pooled leg muscles revealed μDys and EGFP expression. (b) The µDys and EGFP bands were normalized to the GAPDH bands and the ratios were quantified. 1. Untreated wild-type. 2. Untreated mdx4cv. 3. Treated mdx4cv—left leg muscles. 4. Treated mdx4cv—right leg muscles. Dystrophin (MANEX1011B; DSHB, AB-2618171), EGFP (Abcam, Ab290); n.d.u: normalized densitometry units.
Fig 5: The phosphorylation of NPM1 at Ser-125 does not affect nuclear colocalization of NPM1 and Tat. 293 cells were co-transfected with GFP-tagged NPM1 WT, S125A or S125D vector along with Flag-tagged Tat vector following the manufacturer’s protocol of Lipofectamine 3000 (Invitrogen). Additional transfected group of NPM1 WT and FLAG-Tat was treated with 1E7-03. At 24 hrs post-transfection, the cells were fixed with 4% paraformaldehyde (PFA), permeabilized with 0.25% Triton X-100, immunoblotted with antibodies against GFP (ab290, Abcam) and FLAG (F1804, Sigma-Aldrich), and incubated with corresponding secondary antibodies conjugated with Alex Fluor 488 and 594 dyes (Thermo Fisher Scientific). Cells were also stained with Hoechst (Thermo Fisher Scientific). Stained cells were photographed on Olympus IX73 using filter for FITC, TXRED and DAPI fluorescence at 600X magnification. Person’s correlation analysis was conducted in ImageJ.
Supplier Page from Abcam for Anti-GFP antibody