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CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus - BSA Free

Novus Biologicals, part of Bio-Techne | Catalog # NBP2-36440

Novus Biologicals, part of Bio-Techne

Key Product Details

Species Reactivity

Validated:

Bacteria

Cited:

Human, Mouse, Bacteria, Bacterial, Fungi

Applications

Validated:

Immunoblotting, Immunocytochemistry/ Immunofluorescence, Immunohistochemistry, Immunohistochemistry Whole-Mount, Immunohistochemistry-Frozen, Immunoprecipitation, Knockout Validated, Simple Western, Western Blot

Cited:

IF/IHC, Immunocytochemistry/ Immunofluorescence, Immunohistochemistry-Frozen, Knockout Validated, Simple Western, Western Blot

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG1 kappa Clone # 7A9-3A3

Format

BSA Free

Concentration

1.0 mg/ml

Product Specifications

Immunogen

This CRISPR-Cas9 antibody (7A9-3A3) - N-Terminus was raised against Recombinant Cas9 within the N-terminal region of Streptococcus pyogene. .

Specificity

This CRISPR-Cas9 antibody (7A9-3A3) - N-Terminus is specific toCas9 protein from Streptococcus pyogene serotype M1.

Clonality

Monoclonal

Host

Mouse

Isotype

IgG1 kappa

Theoretical MW

158.4 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus - BSA Free

Simple Western: CRISPR-Cas9 Antibody (7A9-3A3)N-TerminusBSA Free [NBP2-36440]

Simple Western: CRISPR-Cas9 Antibody (7A9-3A3)N-TerminusBSA Free [NBP2-36440]

Simple Western: CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus [NBP2-36440] - Image shows a specific band for Cas9 (observed molecular weight ~158 kDa) in HeLa Cas9 lysate but not in Hela WT lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.
Immunocytochemistry/ Immunofluorescence: CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus - BSA Free [NBP2-36440]

Immunocytochemistry/ Immunofluorescence: CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus - BSA Free [NBP2-36440]

Immunocytochemistry/Immunofluorescence: CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus [NBP2-36440] - Hela cells were transiently transfected with an N-terminally Flag-tagged S. pyogenes Cas9 expression vector. The cells were stained with the Cas9 antibody followed by anti mouse-AF488 coupled secondary antibody. Nuclei were counter-stained with Hoechst 33342.
Immunoprecipitation: CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus - BSA Free [NBP2-36440]

Immunoprecipitation: CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus - BSA Free [NBP2-36440]

Immunoprecipitation: CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus [NBP2-36440] - HEK293T expressing N-terminally Flag-tagged S.pyogenes Cas9 were lysed 72h post transfection by resuspending the cells in Hunt buffer and subjecting to 3 freeze-thaw cycles in liquid nitrogen/ice. Proteins were immunoprecipitated from 100ug of whole cell lysate for 1h at 4C with Cas9 supernatant followed by incubation for 1h at 4C with a 1:1 mixture of protein A/G sepharose beads, or for 2h at 4C with Cas9 ab crosslinked to a 1:1 mixture of protein A/G sepharose beads. Beads were washed 2x with Hunt buffer and 1x with TBS. Bound proteins were eluted by boiling in Laemmli, separated by SDS-PAGE and transferred to nitrocellulose. Membrane was blocked, incubated with Cas9 ab, incubated with HRP anti-mouse secondary. *IgG heavy chain

Applications for CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus - BSA Free

Application
Recommended Usage

Immunoblotting

reported in scientific literature (PMID 31959836)

Immunocytochemistry/ Immunofluorescence

1:500

Immunohistochemistry-Frozen

reported in scientific literature (PMID 28153089)

Knockout Validated

reported in scientific literature (PMID 31959836)

Western Blot

1:1000
Application Notes
IF and IHC use of CRISPR-Cas9 antibody (clone 7A9-3A3) on 4% formaldehyde fixed and 20um thick frozen-/cryo-sections reported in scientific literature

Reviewed Applications

Read 8 reviews rated 4.6 using NBP2-36440 in the following applications:

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

PBS

Format

BSA Free

Preservative

0.02% Sodium Azide

Concentration

1.0 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: CRISPR-Cas9

Clustered regularly interspaced short palindromic repeats (CRISPRs) are derived from DNA fragments of bacteriophages that infect prokaryotes. When infected, the bacteria capture snips of DNA from the invading virus to create CRISPR arrays. During subsequent infections, the bacteria produce RNA segments from the CRISPR arrays to target the virus' DNA. CRISPR-associated protein 9 (Cas9) is RNA-guided, binds DNA, and is a cleaving enzyme that functions as an integral component of the bacterial CRISPR adaptive immune system that targets the virus' DNA to disable it (1). To check for sites complementary to the 20 base pair spacer region of the guide RNA (gRNA) of the CRISPR, Cas9 unwinds foreign DNA that invades the bacteria. If the DNA substrate is complementary to the gRNA, Cas9 cleaves the invading DNA, rendering the virus disabled. The presence of a 5'-NGG-3' protospacer adjacent motif (PAM) sequence immediately downstream of the target DNA (protospacer) is required for Cas9 cleavage of foreign DNA. As PAM is absent in bacterial CRISPR loci, cleavage of the host genome is avoided and provides a novel sequence for identification of foreign DNA by Cas9.

Using CRISPR-Cas9 technology, double-stranded DNA breaks may be induced within specific targeted genome sequences (target DNA; protospacer) for insertion or removal of DNA sequences for gene editing applications. To target a specific loci, a gRNA that will bind to a specific target sequence of DNA within a genome is created. The gRNA will recognize the DNA sequence, and the Cas9 enzyme will cleave the DNA at the targeted location. Once the targeted DNA is removed by Cas9, the cell's own DNA repair mechanism is used to insert or remove a DNA sequence for genomic editing.

Cas9 detection is used to confirm and evaluate CRISPR Cas9 gRNA transfection efficiency. Western blot analysis of CRISPR-Cas9 gRNA transfected cell lysates with Cas9 antibodies identifies the protein having a theoretical molecular weight of 160kDa. Broad areas of research are benefiting from CRISPR-Cas9 based gene editing tools including studies of basic immunity functions, genetic screening and disease treatment (2). Ethical concerns have led to many countries making it illegal to manipulate human germline cells or perform embryo genome editing.

References

1. Oakes, B. L., Fellmann, C., Rishi, H., Taylor, K. L., Ren, S. M., Nadler, D. C., . . . Savage, D. F. (2019). CRISPR-Cas9 Circular Permutants as Programmable Scaffolds for Genome Modification. Cell, 176(1-2), 254-267.e216. doi:10.1016/j.cell.2018.11.052

2. Chiou, S. H., Winters, I. P., Wang, J., Naranjo, S., Dudgeon, C., Tamburini, F. B., . . . Winslow, M. M. (2015). Pancreatic cancer modeling using retrograde viral vector delivery and in vivo CRISPR/Cas9-mediated somatic genome editing. Genes Dev, 29(14), 1576-1585. doi:10.1101/gad.264861.115

Long Name

CRISPR-associated Protein 9

Alternate Names

Cas9, CRISPR-associated endonuclease Cas9/Csn1, CRISPR-Cas9/Csn1, CRISPR/Cas9, csn1, SPy_1046, SPy1046, SpyCas9

Additional CRISPR-Cas9 Products

Product Documents for CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus - BSA Free

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot number in the search box below.

Product Specific Notices for CRISPR-Cas9 Antibody (7A9-3A3) - N-Terminus - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

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