The CRISPR-Cas9 system has proven to be a powerful tool for genome editing, allowing for the precise modification of specific DNA sequences within a cell. Many efforts are currently underway to use the CRISPR-Cas9 system for the therapeutic correction of human genetic diseases. The best characterized CRISPR-associated nucleases are the Cas9 proteins from Streptococcus pyogenes (S. pyogenes) and Staphylococcus aureus (S. aureus).

Check the success of transfection

Detect Cas9 protein expression level by western blot with anti-Cas9 antibodies, this can indicate whether the transfection is successful.

Check the Cas9 protein was delivered to the nucleus

CRISPR-Cas9 must translocate to the nuclei of the transfected cells for executing its nuclease activity on the genomic DNA. Cas9 protein must be checked through ICC/IF or IHC staining of the transfected cells with CRISPR-Cas9 antibodies.

Check the CRISPR-Cas9 expression levels

The level of the duration of expression of Cas9 protein is very critical when using CRISPR-Cas9 genome editing technology. In stable clones, it is suggested to isolate multiple clones and to screen them for Cas9 expression levels through western blot analysis. In Transient systems, check that Cas9 expression was transient by using western blot and anti-Cas9 antibodies.

Confirming the binding specificity of CRISPR-Cas9

CRISPR-Cas9 genome editing system has two major components: a synthetic RNA “guide RNA” (gRNA) and a non-specific CRISPR-Cas9 protein with nuclease activity. Structurally, the gRNA is composed of a “scaffold” sequence which is essential for Cas9 enzyme’s binding to gRNA and a user-designated “spacer” or “targeting” sequence which defines the genomic target to be manipulated. Using ChIP grade CRISPR-Cas9 antibodies, it is very easy to detect/quantify the binding specificity of Cas9 enzyme with a given gRNA and targeted or non-targeted region primers.

Creative Diagnostics, a dedicated supplier of high quality Cas9 antibodies, Creative Diagnostics now offers the largest collection of optimized anti-CRISPR/Cas9 antibodies, validated in a number of different applications including western blot (WB), immunofluorescence (IF), immunoprecipitation (IP), and chromatin immunoprecipitation (ChIP). Creative Diagnostics offers both monoclonal and polyclonal antibodies raised against recombinant Cas9 protein from Streptococcus pyogenes (S. pyogenes).

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Creative Diagnostics is a leading manufacturer and supplier of antibodies, viral antigens, innovative diagnostic components and critical assay reagents. We provide contract biologic R&D and manufacturing services to the diagnostic manufacturers along with GMP biologics manufacturing for the biopharmaceutical market. Our goal is to provide a trusted source for all your assay development and manufacturing needs.