![GBA was immunoprecipitated using 0.5mg Hela whole cell extract, 5µg of Rabbit polyclonal to GBA and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).The antibody was incubated under agitation with Protein G beads for 10min, Hela whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab92997.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 60kDa; GBA](http://www.bioprodhub.com/system/product_images/ab_products/2/sub_2/23158_ab92997-171828-IPV018ab9299720min.jpg)
GBA was immunoprecipitated using 0.5mg Hela whole cell extract, 5µg of Rabbit polyclonal to GBA and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).The antibody was incubated under agitation with Protein G beads for 10min, Hela whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab92997.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 60kDa; GBA
All lanes : Anti-GBA antibody (ab92997) at 1 µg/mlLane 1 : MCF7 (Human breast adenocarcinoma cell line) Whole Cell LysateLane 2 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate Lane 3 : U2OS (Human osteosarcoma cell line) Whole Cell LysateLane 4 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate Lysates/proteins at 10 µg per lane.SecondaryGoat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilutiondeveloped using the ECL techniquePerformed under reducing conditions.
ICC/IF image of ab92997 stained HepG2 cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab92997 at 5µg/ml overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti- rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in Methanol (100%, 5 minutes) fixed HeLa, Hek293, HepG2, and MCF-7 cell lines at 5ug/ml.