Guide-itTM Cas9 Monoclonal Antibody (Clone TG8C1)

Description

Takara Bio offers both monoclonal and polyclonal antibodies raised against recombinant Cas9 protein fromStreptococcus pyogenes. Our products are to be used forResearch Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval. Back The Guide-it Cas9 Monoclonal Antibody Clone TG8C1 , which was raised in mouse against recombinant full-length Cas9 rCas9 , was tested via Western blot analysis using either increasing concentrations of recombinant Cas9 Panel A or lysates made from cells transiently expressing Cas9 after plasmid transfection Panel B . Panel A: Increasing concentrations of rCas9 1 ng, 2 ng, 3 ng, and 5 ng were analyzed via Western blotting. The samples were separated using a 7.5 % SDS PAGE gel, followed by transfer onto a membrane. The membrane was probed with a 1:3,000 dilution of the monoclonal anti-Cas9 antibody and detection was performed using an anti-mouse HRP-coupled secondary antibody at a 1:10,000 dilution. A clearly visible band at the expected molecular weight of 160 kDa was detected in each lane containing rCas9, even for the smallest amount of rCas9 loaded 1 ng . No signal was detected in the HEK 293 lane, which contains a lysate of 5,000 untransfected HEK 293 cells. Panel B: HEK 293 cells were transiently transfected with a plasmid encoding Cas9 protein. Lysates made from an increasing number of Cas9-expressing cells 1,000, 2,000, and 5,000 cells were separated using a 7.5 % SDS PAGE gel, followed by transfer onto a membrane. The membrane was probed with a 1:3,000 dilution of the monoclonal anti-Cas9 antibody and detection was performed using an anti-mouse HRP-coupled secondary antibody at a 1:10,000 dilution. A clearly visible band at 160 kDa was detected only in the lanes containing lysates of transfected cells. No signal was detected in the HEK 293 lane, which contains a lysate of untransfected HEK 293 cells, and the NIH 3T3 lane, which contains a lysate of untransfected NIH 3T3 cells. The Marker lanes contain a molecular weight marker. Back The Cas9 monoclonal antibody detects Cas9 protein by immunocytochemistry ICC .RPE cells stably expressing ZsGreen1 were transfected with 0.35 mug of a plasmid encoding wild-type Cas9 under the control of the SV40 promoter using Xfect Transfection Reagent. Cells were fixed with 4 % paraformaldehyde 48 hours post-transfection and stained with Cas9 monoclonal antibody 1:100 for one hour at room temperature RT , followed by staining with donkey anti-mouse secondary antibody 1:1,000 for 30 minutes at RT. The antifade mounting media contained DAPI, allowing for nuclear staining.