CRISPR/Cas9 Antibody (C15200229)

Lot	002
Concentration	2 μg/μl
Species reactivity	Streptococcus pyogenes
Type	Monoclonal
Purity	Protein G purified
Host	Mouse
Precautions	This product is for research use only. Not for use in diagnostic or therapeutic procedures.

Applications	Suggested dilution	References
ChIP ^*	5 μg/ChIP	Fig 1
Western Blotting	1:1,000 - 1:10,000	Fig 2
Immunoprecipitation	6 μg/IP	Fig 3
Immunofluorescence	1:400	Fig 4

CRISPR/Cas9 Antibody for ChIP

Figure 1. ChIP using the Diagenode monoclonal antibody directed against Cas9
ChIP was performed on NIH3T3 cells stably expressing GFP-H2B, nuclease dead Cas9, and a GFP-targeting gRNA. 50 μg chromatin was incubated overnight at 4°C with 5 or 10 μg of either an anti-FLAG antibody or the Diagenode antibody against Cas9 (Cat. No. C15200229). Mouse IgG was used as a negative IP control. qPCR was performed with primers specific for the GFP gene, and for a non-targeted region (protein kinase C delta, Prkcd), used as negative control. Figure 1 shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).

CRISPR/Cas9 Antibody for Western Blot

Figure 2. Western blot analysis using the Diagenode monoclonal antibody directed against CRISPR/Cas9
Figure 1A: Western blot was performed on protein extracts from HeLa cells transfected with Cas9 using the Diagenode antibody against CRISPR/Cas9 (Cat. No. C15200229). The antibody was used at different dilutions. The marker is shown on the left, position of the Cas9 protein is indicated on the right. Figure 1B: Western blot was performed on protein extracts from HeLa cells transfected with Cas9 (lane 1) or from untransfected cells (lane 2) using the Diagenode antibody against CRISPR/Cas9 (Cat. No. C15200229), diluted 1:4,000 in PBS-T containing 3% NFDM. The marker is shown on the left, position of the Cas9 protein is indicated on the right.

CRISPR/Cas9 Antibody for Immunoprecipitation

Figure 3. IP using the Diagenode monoclonal antibody directed against Cas9
IP was performed on whole cell extracts (300 μg) from HEK293 cells transfected with a Cas9 expression vector (lane 1 and 3), or untransfected cells (lane 2 and 4) using 6 μg of the Diagenode antibody against Cas9 (Cat. No. C15200229). The immunoprecipitated proteins were subsequently analysed by Western blot with the polyclonal Cas9 antibody (Cat. No. C15310258, diluted 1:8,000). Lane 3 and 4 show the result of the IP, the input (15 μg) is shown in lane 1 and 2.

CRISPR/Cas9 Antibody for Immunofluorescence

Figure 4. Immunofluorescence using the Diagenode monoclonal antibody directed against CRISPR/Cas9
HeLa cells transfected with a Cas9 expression vector (left) or untransfected cells (right) were fixed in methanol at -20°C, permeabilized with acetone at -20°C and blocked with PBS containing 2% BSA. The cells were stained with the Cas9 N-terminal antibody (Cat. No. C15200229) diluted 1:400, followed by incubation with an anti-mouse secondary antibody coupled to AF488. The bottom images show counter-staining of the nuclei with Hoechst 33342.

CRISPR systems are adaptable immune mechanisms which are present in many bacteria to protect themselves from foreign nucleic acids, such as viruses, transposable elements or plasmids. Recently, the CRISPR/Cas9 (CRISPR-associated protein 9 nuclease, UniProtKB/Swiss-Prot entry Q99ZW2) system from S. pyogenes has been adapted for inducing sequence-specific double stranded breaks and targeted genome editing. This system is unique and flexible due to its dependence on RNA as the moiety that targets the nuclease to a desired DNA sequence and can be used to induce indel mutations, specific sequence replacements or insertions and large deletions or genomic rearrangements at any desired location in the genome. In addition, Cas9 can also be used to mediate upregulation of specific endogenous genes or to alter histone modifications or DNA methylation.

WB
Western blot : The quality of antibodies used in this technique is crucial for correct and specific protein identification. Diagenode offers huge selection of highly sensitive and specific western blot-validated antibodies. Learn more about: Load... Read more

IF
Immunofluorescence: Diagenode offers huge selection of highly sensitive antibodies validated in IF. Immunofluorescence using the Diagenode monoclonal antibody directed against CRISPR/Cas9 HeLa cells transfected with a Cas9 expression vector (... Read more

IP
Immunoprecipitation Read more

ChIP-qPCR (ab)
Read more

CRISPR/Cas9 monoclonal antibody C15200229 DATASHEET CRISPR/Cas9 monoclonal antibody datasheet	Download
Epigenetic Antibodies Brochure BROCHURE More than in any other immuoprecipitation assays, quality antibodies are critical tools in many e...	Download
Antibodies you can trust POSTER Epigenetic research tools have evolved over time from endpoint PCR to qPCR to the analyses of lar...	Download

CRISPR/Cas9 Antibody SDS US en	Download
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TurboCas: A method for locus-specific labeling of genomic regions and isolating their associated protein interactome
Bercin K Cenik et al.
Regulation of gene expression during development and stress response requires the concerted action of transcription factors and chromatin-binding proteins. Because this process is cell-type specific and varies with cellular conditions, mapping of chromatin factors at individual regulatory loci is crucial for underst...

Genome-wide CRISPR screen identifies protein pathways modulating tauprotein levels in neurons
Sanchez C. G. et al.
Aggregates of hyperphosphorylated tau protein are a pathological hallmark of more than 20 distinct neurodegenerative diseases, including Alzheimer’s disease, progressive supranuclear palsy, and frontotemporal dementia. While the exact mechanism of tau aggregation is unknown, the accumulation of aggregates corr...

Transgenic mice for in vivo epigenome editing with CRISPR-based systems
Gemberling, M. et al.
The discovery, characterization, and adaptation of the RNA-guided clustered regularly interspersed short palindromic repeat (CRISPR)-Cas9 system has greatly increased the ease with which genome and epigenome editing can be performed. Fusion of chromatin-modifying domains to the nuclease-deactivated form of Cas9 (dCa...

Circuit-specific hippocampal ΔFosB underlies resilience to stress-induced social avoidance.
Eagle, Andrew L and Manning, Claire E and Williams, Elizabeth S and Bastle, Ryan M and Gajewski, Paula A and Garrison, Amber and Wirtz, Alexis J and Akguen, Seda and Brandel-Ankrapp, Katie and Endege, Wilson and Boyce, Frederick M and Ohnishi, Yoshinori N
Chronic stress is a key risk factor for mood disorders like depression, but the stress-induced changes in brain circuit function and gene expression underlying depression symptoms are not completely understood, hindering development of novel treatments. Because of its projections to brain regions regulating reward a...

Endogenous retroviruses are a source of enhancers with oncogenic potential in acute myeloid leukaemia
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Acute myeloid leukemia (AML) is a highly aggressive hematopoietic malignancy, defined by a series of genetic and epigenetic alterations, which result in deregulation of transcriptional networks. One understudied but important source of transcriptional regulators are transposable elements (TEs), which are widespread ...

Improved double-nicking strategies for COL7A1 editing by homologous recombination
Kocher Thomas, Wagner Roland N., Klausegger Alfred, Guttmann-Gruber Christina, Hainzl Stefan, Bauer Johann W., Reichelt Julia, Koller Ulrich
Current gene editing approaches for treatment of recessive dystrophic epidermolysis bullosa (RDEB), an inherited, severe form of blistering skin disease, suffer from low efficiencies and safety concerns that complicate implementation in clinical settings. We present a strategy for efficient and precise repair of RDE...

Improved Double-Nicking Strategies for COL7A1-Editing by Homologous Recombination.
Kocher T, Wagner RN, Klausegger A, Guttmann-Gruber C, Hainzl S, Bauer JW, Reichelt J, Koller U
Current gene-editing approaches for treatment of recessive dystrophic epidermolysis bullosa (RDEB), an inherited, severe form of blistering skin disease, suffer from low efficiencies and safety concerns that complicate implementation in clinical settings. We present a strategy for efficient and precise repair of RDE...

Essential Gene Profiles for Human Pluripotent Stem Cells Identify Uncharacterized Genes and Substrate Dependencies.
Mair B, Tomic J, Masud SN, Tonge P, Weiss A, Usaj M, Tong AHY, Kwan JJ, Brown KR, Titus E, Atkins M, Chan KSK, Munsie L, Habsid A, Han H, Kennedy M, Cohen B, Keller G, Moffat J
Human pluripotent stem cells (hPSCs) provide an invaluable tool for modeling diseases and hold promise for regenerative medicine. For understanding pluripotency and lineage differentiation mechanisms, a critical first step involves systematically cataloging essential genes (EGs) that are indispensable for hPSC fitne...

RNA-Based dCas9–VP64 System Improves the Viability of Cryopreserved Mammalian Cells
Hu Yong, Li Lei, Yu Yin, Huang Haishui, Uygun Basak E., Yarmush Martin L.
Regenerative therapies require availability of an abundant healthy cell source which can be achieved by e±cient cryopreservation techniques. Here, we established a novel approach for improved cell cryopreservation using an mRNA-based dCas9-VP64 gene activation system for transient, yet highly e±cient e...

CRISPR-Mediated Gene Targeting of Human Induced Pluripotent Stem Cells
Susan M. Byrne, George M. Church
CRISPR/Cas9 nuclease systems can create double-stranded DNA breaks at specific sequences to efficiently and precisely disrupt, excise, mutate, insert, or replace genes. However, human embryonic stem cells and induced pluripotent stem cells (iPSCs) are more difficult to transfect and less resilient to DNA damage than...

CRISPR/Cas9 Antibody (sample size)

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