Recombinant
RabMAb

Recombinant Anti-CRISPR-Cas9 antibody [EPR19799] (ab203933)

Overview

  • Product name

    Anti-CRISPR-Cas9 antibody [EPR19799]
    See all CRISPR-Cas9 primary antibodies
  • Description

    Rabbit monoclonal [EPR19799] to CRISPR-Cas9
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, ICC/IF, Flow Cyt, IP, IHC-P, ICCmore details
  • Species reactivity

    Reacts with: Staphylococcus aureus
  • Immunogen

    Synthetic peptide within CRISPR-Cas9 aa 100-200. The exact sequence is proprietary. (Staphylococcus aureus subsp. Aureus).
    Database link: J7RUA5

  • Positive control

    • WB: HEK-293 whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, S.aureus) with Myc-His tag. ICC: HeLa cells transfected with S.aureus Cas9 (pcDNA3.1(+)-Myc-His). ICC/IF: 293T cells transfected with CRISPR-Cas9 (J7RUA5, S.aureus) with Myc-His tag. Flow Cyt: HEK-293 whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, S.aureus) with Myc-His tag. IP: HEK-293 whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, S.aureus) with Myc-His tag.
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    This product is a recombinant rabbit monoclonal antibody.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
  • Storage buffer

    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR19799
  • Isotype

    IgG

Applications

Our Abpromise guarantee covers the use of ab203933 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
WB 1/20000. Detects a band of approximately 124 kDa (predicted molecular weight: 124 kDa).
ICC/IF 1/500.
Flow Cyt 1/60.
IP 1/30.
IHC-P 1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC 1/500.

Target

  • Relevance

    [FUNCTION] CRISPR (clustered regularly interspaced short palindromic repeat) is an adaptive immune system that provides protection against mobile genetic elements (viruses, transposable elements and conjugative plasmids). CRISPR clusters contain spacers, sequences complementary to antecedent mobile elements, and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA) (Probable). In type II CRISPR systems correct processing of pre-crRNA requires a trans-encoded small RNA (tracrRNA), endogenous ribonuclease 3 (rnc) and this protein. The tracrRNA serves as a guide for ribonuclease 3-aided processing of pre-crRNA. Subsequently Cas9/crRNA/tracrRNA endonucleolytically cleaves linear or circular dsDNA target complementary to the spacer. The target strand not complementary to crRNA is first cut endonucleolytically, then trimmed by 3'-5' exonucleolytically. DNA-binding requires protein and both RNA species. Cas9 probably recognizes a short motif in the CRISPR repeat sequences (the PAM or protospacer adjacent motif) to help distinguish self versus nonself.
  • Database links

    • Alternative names

      • Cas9 antibody
      • CRISPR-associated endonuclease Cas9/Csn1 antibody
      • CRISPR-Cas9/Csn1 antibody
      • csn1 antibody
      • SpyCas9 antibody
      see all

    Images

    • All lanes : Anti-CRISPR-Cas9 antibody [EPR19799] (ab203933) at 1/20000 dilution

      Lane 1 : Empty vector with GFP-Myc tag (vector control) transfected HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
      Lane 2 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (Q99ZW2, Streptococcus pyogenes serotype M1) with GFP-Myc tag
      Lane 3 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus, N–terminal aa1-800) with GFP-Myc tag
      Lane 4 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus, C–terminal aa801-1409) with GFP-Myc tag
      Lane 5 : Empty vector with Myc-His tag (vector control) transfected HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
      Lane 6 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (A1IQ68, Neisseria meningitidis serogroup A / serotype 4A (strain Z2491)) with Myc-His tag
      Lane 7 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus) with Myc-His tag
      Lane 8 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (Q03JI6, Streptococcus thermophilus (strain ATCC BAA-491 / LMD-9)) with Myc-His tag
      Lane 9 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

      Predicted band size: 124 kDa
      Observed band size: 124 kDa


      Exposure time: 1 second


      Blocking/Dilution buffer: 5% NFDM/TBST.

    • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized 293T (Human epithelial cell line from embryonic kidney) cells transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag or Empty vector, labeling CRISPR-Cas9 with ab203933 at 1/500 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

      Confocal image showing positive staining on 293T cells transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag. 

      The nuclear counterstain is DAPI (blue).

      Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

      The negative controls are as follows:-

      -ve control 1: ab203933 at 1/500 dilution followed by ab150120  at 1/1000 dilution.

      -ve control 2: ab7291  at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

    • All lanes : Anti-CRISPR-Cas9 antibody [EPR19799] (ab203933) at 1/20000 dilution

      Lane 1 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag
      Lane 2 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
      Lane 3 : NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate
      Lane 4 : Rat embryo lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

      Predicted band size: 124 kDa
      Observed band size: 124 kDa


      Exposure time: 5 seconds


      Blocking/Dilution buffer: 5% NFDM/TBST.

    • Immunocytochemical analysis of agarose-embedded 293T (Human epithelial cell line from embryonic kidney) cells transfected with Staphylococcus aureus subsp. Aureus Cas9 (pcDNA3.1(+)-Myc-His) labeling CRISPR-Cas9 with ab203933 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. 

      Positive staining on 293T cells transfected with Staphylococcus aureus subsp. Aureus Cas9 (pcDNA3.1(+)-Myc-His) is observed.

      Counter stained with Hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

    • Flow cytometric analysis of 4% paraformaldehyde-fixed HEK-293 (Human epithelial cell line from embryonic kidney) cells transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag labeling CRISPR-Cas9 with ab203933 at 1/60 dilution (red) compared with a Rabbit IgG,monoclonal [EPR25A] -Isotype control (ab172730) (black). Goat anti Rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

    • CRISPR-Cas9 was immunoprecipitated from 0.35 mg of HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag with ab203933 at 1/30 dilution.

      Western blot was performed from the immunoprecipitate using ab203933 at 1/1000 dilution.

      VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

      Lane 1: HEK-293 whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag 10µg (Input).

      Lane 2: ab203933 IP in HEK-293 whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag.

      Lane 3: Rabbit IgG,monoclonal [EPR25A] - Isotype Control (ab172730) instead of ab203933 in HEK-293 whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag.

      Blocking and dilution buffer and concentration: 5% NFDM/TBST.

      Exposure time: 1 second.

    • Immunohistochemical analysis of paraffin-embedded Human endometrium tissue labeling CRISPR-Cas9 with ab203933 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. 

      No staining on Human endometrium is observed.

      Counter stained with Hematoxylin.

    • Immunocytochemical analysis of agarose-embedded HeLa (Human epithelial cell line from cervix adenocarcinoma) cells transfected with blank pcDNA3.1(+)-Myc-His vector labeling CRISPR-Cas9 with ab203933 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
      No staining on HeLa cells transfected with blank pcDNA3.1(+)-Myc-His vector.
      Counter stained with Hematoxylin.

       

    • Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling CRISPR-Cas9 with ab203933 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. 

      No staining on mouse spleen is observed.

      Counter stained with Hematoxylin.

    • Immunohistochemical analysis of paraffin-embedded Rat spleen tissue labeling CRISPR-Cas9 with ab203933 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. 

      No staining on rat spleen is observed.

      Counter stained with Hematoxylin.

    References

    ab203933 has not yet been referenced specifically in any publications.

    Customer reviews and Q&As

    Application
    Immunohistochemistry (Frozen sections)
    Sample
    Mouse Tissue sections (spinal cord, hSyn-SaCas9 delivered by AAV)
    Permeabilization
    Yes - 0.3% TritonX-100
    Specification
    spinal cord, hSyn-SaCas9 delivered by AAV
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 20°C
    Fixative
    Paraformaldehyde

    Abcam user community

    Verified customer

    Submitted Feb 06 2019

    Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
    For licensing inquiries, please contact partnerships@abcam.com

    Sign up