Goat anti human igg fitc

PANC-1 cells were cultured in a Lab-Tek chamber slide (Nalge Nunc International, Rochester, NY, USA) at a density of 20,000 cells/well. After 48 h, cells were fixed with 4% paraformaldehyde in phosphate buffered saline (PBS) for 15 min. After being permeabilized with 0.5% Triton X-100 in PBS for 10 min, the cells were blocked with 1% BSA in PBS and incubated with primary antibodies overnight at 4 °C. Primary antibodies used in these studies were anti-pericentrin (Abcam, Cambridge, UK), anti-α-tubulin (Abcam), human anti-CREST (Immuno Vision Inc., Springdale, AR, USA) and anti-cleaved caspase-3 (1:400, Cell Signaling Technology). The cells were then washed three times with PBS, and incubated with the indicated secondary antibody for 2 h at 25 °C. Secondary antibodies were goat Alexa Fluor 568 (Invitrogen), goat Alexa Fluor 488 (Abcam), and goat anti-Human IgG-FITC (Invitrogen). Nuclei were counterstained with DAPI and mounted with ProLong Gold Antifade (Invitrogen). Images were captured using a ZEISS LSM 710 confocal microscope and processed using ZEN software (ZEISS International, Oberkochen, DE).

Briefly, 293 T cells were respectively transfected with HIV-1_BJMSM2316 and HIV-1_{BJMSM2316_D368R} GP160 expressing plasmids and harvested after 48 h. Cells were incubated with F6 (10 μg/ml) or other control nAbs (VRC01 and PG16) at 37 °C for 30 min and then detected by 1:1000 diluted goat anti-human IgG-FITC (Invitrogen). Flow cytometric data were acquired on an LSR Fortessa flow cytometer (BD Biosciences) and analyzed with FlowJo software (TreeStar). Statistical analyses were performed with a t test using the software GraphPad Prism 5.01. P < 0.05 was considered significant.