Recombinant Anti-CRISPR-Cas9 antibody [EPR18991] (ab189380)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR18991] to CRISPR-Cas9
- Suitable for: ICC, IHC-P, ICC/IF, WB, Flow Cyt (Intra)
Related conjugates and formulations
Overview
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Product name
Anti-CRISPR-Cas9 antibody [EPR18991]
See all CRISPR-Cas9 primary antibodies -
Description
Rabbit monoclonal [EPR18991] to CRISPR-Cas9 -
Host species
Rabbit -
Tested applications
Suitable for: ICC, IHC-P, ICC/IF, WB, Flow Cyt (Intra)more details -
Species reactivity
Predicted to work with: Streptococcus pyogenes -
Immunogen
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB and Flow Cyt (intra): HEK-293 whole cell lysate transfected with CRISPR-Cas9 (Q99ZW2, Streptococcus pyogenes serotype M1) with GFP-Myc tag. IHC: 293T cells transfected with Streptococcus pyogenes serotype M1 Cas9 (pcDNA3.1(+)-GFP-Myc). ICC/IF: 293T cells transfected with CRISPR-Cas9 with GFP-tag.
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General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
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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
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol, 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR18991 -
Isotype
IgG
Associated products
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Alternative Versions
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Compatible Secondaries
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Conjugation kits
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Isotype control
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab189380 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 |
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ICC |
1/100.
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IHC-P |
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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ICC/IF |
1/500.
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WB | (1) |
1/20000. Detects a band of approximately 184 kDa (predicted molecular weight: 158 kDa).
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Flow Cyt (Intra) |
1/70.
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Notes |
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ICC
1/100. |
IHC-P
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
ICC/IF
1/500. |
WB
1/20000. Detects a band of approximately 184 kDa (predicted molecular weight: 158 kDa). |
Flow Cyt (Intra)
1/70. |
Target
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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
- Entrez Gene: 901176 Streptococcus pyogenes
- SwissProt: Q99ZW2 Streptococcus pyogenes
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Alternative names
- Cas9 antibody
- CRISPR-associated endonuclease Cas9/Csn1 antibody
- CRISPR-Cas9/Csn1 antibody
see all
Images
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All lanes : Anti-CRISPR-Cas9 antibody [EPR18991] (ab189380) at 1/20000 dilution
Lane 1 : 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 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: 158 kDa
Observed band size: 184 kDa why is the actual band size different from the predicted?
Exposure time: 10 secondsBlocking/Dilution buffer: 5% NFDM/TBST.
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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-GFP or GFP only, labeling CRISPR-Cas9 with ab189380 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 647) (ab150079) secondary antibody at 1/1000 dilution.
Confocal image showing positive staining on 293T cells transfected with CRISPR-Cas9 with GFP-tag.
The nuclear counterstain is DAPI (blue).
Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (Alexa Fluor® 594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows:-
-ve control 1: ab189380 at 1/500 dilution followed by ab150120 (Alexa Fluor® 594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150079 (Alexa Fluor® 647 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.
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Immunohistochemical analysis of paraffin-embedded Rat cerebrum tissue labeling CRISPR-Cas9 with ab189380 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
No staining on rat cerebrum is observed.
Counter stained with Hematoxylin.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Intracellular Flow Cytometry analysis of 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) labelling CRISPR-Cas9 (red) with ab189380 at dilution of 1/70. The secondary antibody used was Alexa Fluorr® 488 goat-anti-rabbit IgG (1/2000). Cells were fixed with 4% paraformaldehyde. Isotype control antibody was (ab172730) Rabbit monoclonal IgG (black).
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Immunocytochemical analysis of agarose-embedded sections of 293T (Human epithelial cell line from embryonic kidney) cells transfected with Streptococcus pyogenes serotype M1 Cas9 (pcDNA3.1(+)-GFP-Myc) labeling CRISPR-Cas9 with ab189380 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
Positive staining on 293T cells transfected with Streptococcus pyogenes serotype M1 Cas9 (pcDNA3.1(+)-GFP-Myc) is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
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All lanes : Anti-CRISPR-Cas9 antibody [EPR18991] (ab189380) 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 : Empty vector with Myc-His tag (vector control) transfected HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
Lane 5 : 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 6 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus) with Myc-His tag
Lane 7 : 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 8 : 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: 158 kDa
Observed band size: 184 kDa why is the actual band size different from the predicted?
Exposure time: 3 secondsBlocking/Dilution buffer: 5% NFDM/TBST.
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Immunocytochemical analysis of agarose-embedded sections of 293T (Human epithelial cell line from embryonic kidney) cells transfected with blank pcDNA3.1(+)-GFP-Myc vector labeling CRISPR-Cas9 with ab189380 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
Negative on 293T cells transfected with blank pcDNA3.1(+)-GFP-Myc vector.
Counter stained with Hematoxylin.
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Immunohistochemical analysis of paraffin-embedded Human stomach tissue labeling CRISPR-Cas9 with ab189380 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
No staining on Human stomach is observed.
Counter stained with Hematoxylin.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Immunohistochemical analysis of paraffin-embedded Mouse stomach tissue labeling CRISPR-Cas9 with ab189380 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
No staining on mouse stomach is observed.
Counter stained with Hematoxylin.
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
Certificate of Compliance
References (7)
ab189380 has been referenced in 7 publications.
- Qiu M et al. Lung-selective mRNA delivery of synthetic lipid nanoparticles for the treatment of pulmonary lymphangioleiomyomatosis. Proc Natl Acad Sci U S A 119:N/A (2022). PubMed: 35173043
- Thompson EM et al. The cytotoxic action of BCI is not dependent on its stated DUSP1 or DUSP6 targets in neuroblastoma cells. FEBS Open Bio 12:1388-1405 (2022). PubMed: 35478300
- Stuart WD et al. CRISPRi-mediated functional analysis of NKX2-1-binding sites in the lung. Commun Biol 4:568 (2021). PubMed: 33980985
- Matsunaga W et al. Cancer Cell-specific Transfection of hCas9 Gene Using Ad5F35 Vector. Anticancer Res 41:3731-3740 (2021). PubMed: 34281831
- McCann JL et al. MagnEdit-interacting factors that recruit DNA-editing enzymes to single base targets. Life Sci Alliance 3:N/A (2020). PubMed: 32094150
- Xu S et al. CAS9 is a genome mutator by directly disrupting DNA-PK dependent DNA repair pathway. Protein Cell 11:352-365 (2020). PubMed: 32170574
- Cui L et al. A CRISPRi screen in E. coli reveals sequence-specific toxicity of dCas9. Nat Commun 9:1912 (2018). PubMed: 29765036