IHC image of CD147 staining in human heart formalin fixed paraffin embedded tissue section, performed on a Leica Bond system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab64616,1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
ab64616 stained SV40LT-SMC cells. The cells were 100% methanol fixed (5 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 ab64616 at 5µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- rabbit (ab96899) IgG (H+L) used at a 1/250 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.
![CD147 was immunoprecipitated using 0.5mg Hela whole cell extract, 5µg of Rabbit polyclonal to CD147 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 ab64616.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 60kDa; CD147](http://www.bioprodhub.com/system/product_images/ab_products/2/sub_1/23383_ab64616-205654-IPVI002ab646168m.jpg)
CD147 was immunoprecipitated using 0.5mg Hela whole cell extract, 5µg of Rabbit polyclonal to CD147 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 ab64616.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 60kDa; CD147
All lanes : Anti-CD147 antibody (ab64616) at 1 µg/mlLane 1 : HeLa Whole Cell Lysate - Hydroxyurea Treated (48hr, 2uM)Lane 2 : HeLa Whole Cell Lysate - Bleomycin Treated (20U/ml)Lane 3 : HeLa Whole Cell Lysate - Staurosporine Treated (48hr, 500nM)Lane 4 : Y79 (Human retinoblastoma cell line) Whole Cell LysateLane 5 : Skeletal Muscle (Mouse) Tissue LysateLane 6 : HeLa (Human epithelial carcinoma cell line) Whole Cell LysateLane 7 : SK N SH (Human neuroblastoma) Whole Cell LysateLysates/proteins at 10 µg per lane.SecondaryGoat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilutionPerformed under reducing conditions.