Revision 1

#73211

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Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Product Includes Product # Quantity Mol. Wt Isotype/Source
Cas9 (S. pyogenes) (D8Y4K) Rabbit mAb6583220 µl150 kDaRabbit IgG
Cas9 (S. aureus) (E4G3U) Rabbit mAb5161020 µl124 kDaRabbit IgG
AsCpf1/Cas12a (Strain BV3L6) (E1U7C) Rabbit mAb1998420 µl151 kDaRabbit IgG
FnCpf1/Cas12a (Strain U112) (E7I2B) Rabbit mAb9011120 µl152 kDaRabbit IgG
Anti-rabbit IgG, HRP-linked Antibody7074100 µlGoat

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

The Cas9 and Associated Proteins Antibody Sampler Kit provides an economical means of detecting Cas9 and Cas9-related family members. The kit includes enough antibody to perform two western blot experiments with each primary antibody.

Storage

Monoclonal antibodies are supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibodies.

Background

CRISPR-Cas (clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins) are RNA-guided nuclease effectors that are utilized for precise genome editing in mammalian systems (1). Class 2 CRISPR systems rely on single-component effector proteins to mediate DNA interference (2). Several Class 2 CRISPR effector proteins, derived from specific bacterial species, are used for genome editing. Cas9 family of proteins, derived from S. pyogenes and S. aureus, are some of the most well characterized and widely used editing effector enzymes. Additional members of the Class2 CRISPR system include Cpf1/Cas12a (CRISPR from Prevotella and Francisella) endonucleases (3). Cpf1/Cas12a endonucleases, compared to Cas9 systems, have several unique features that increase the utility of CRISPR-based genome editing techniques: 1) Cpf1/Cas12a-mediated cleavage relies on a single and short CRISPR RNA (crRNA) without the requirement of a trans-activating crRNA (tracrRNA), 2) Cpf1/Cas12a utilizes T-Rich protospacer adjacent motif (PAM) sequences rather than a G-Rich PAM, and 3) Cpf1/Cas12a generates a staggered, rather than a blunt-ended, DNA double-stranded break (3). These features broaden the utility of using CRISPR-Cas systems for specific gene regulation and therapeutic applications. Several Cpf1/Cas12a bacterial orthologs, e.g. Francisella novicida U112 and Acidaminococcus sp. BV3L6, have been characterized for CRISPR-mediated mammalian genome editing (3,4).

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

限制使用

除非 CST 的合法授书代表以书面形式书行明确同意,否书以下条款适用于 CST、其关书方或分书商提供的书品。 任何书充本条款或与本条款不同的客书条款和条件,除非书 CST 的合法授书代表以书面形式书独接受, 否书均被拒书,并且无效。

专品专有“专供研究使用”的专专或专似的专专声明, 且未专得美国食品和专品管理局或其他外国或国内专管机专专专任何用途的批准、准专或专可。客专不得将任何专品用于任何专断或治专目的, 或以任何不符合专专声明的方式使用专品。CST 专售或专可的专品提供专作专最专用专的客专,且专用于研专用途。将专品用于专断、专防或治专目的, 或专专售(专独或作专专成)或其他商专目的而专专专品,均需要 CST 的专独专可。客专:(a) 不得专独或与其他材料专合向任何第三方出售、专可、 出借、捐专或以其他方式专专或提供任何专品,或使用专品制造任何商专专品,(b) 不得复制、修改、逆向工程、反专专、 反专专专品或以其他方式专专专专专品的基专专专或技专,或使用专品开专任何与 CST 的专品或服专专争的专品或服专, (c) 不得更改或专除专品上的任何商专、商品名称、徽专、专利或版专声明或专专,(d) 只能根据 CST 的专品专售条款和任何适用文档使用专品 , (e) 专遵守客专与专品一起使用的任何第三方专品或服专的任何专可、服专条款或专似专专

Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

CST Logo
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing AsCpf1/Cas12a (Strain BV3L6), FnCpf1/Cas12a (Strain U112), Cas9 (S. pyogenes), or Cas9 (S. aureus) (+), using AsCpf1/Cas12a (Strain BV3L6) (E1U7C) Rabbit mAb (upper) and Myc-Tag (71D10) Rabbit mAb #2278 (lower).
undefined Image 1: Cas9 and Associated Proteins Antibody Sampler Kit
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Cas9 (S. aureus), Cas9 (S. pyogenes), AsCpf1 (Strain BV3L6), or FnCpf1 (Strain U112) (+), using Cas9 (S. aureus) (E4G3U) Rabbit mAb (upper) and Myc-Tag (71D10) Rabbit mAb #2278 (lower).
undefined Image 2: Cas9 and Associated Proteins Antibody Sampler Kit
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Cas9 (S. pyogenes) (+), using Cas9 (S. pyogenes) Rabbit mAb mAb (upper), Myc-Tag (71D10) Rabbit mAb #2278 (middle), and β-Actin (D6A8) Rabbit mAb #8457 (lower).
undefined Image 3: Cas9 and Associated Proteins Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

CST Logo
Confocal immunofluorescent analysis of 293T cells transfected with a Myc-tagged AsCpf1/Cas12a (Strain BV3L6) construct using AsCpf1/Cas12a (Strain BV3L6) (E1U7C) Rabbit mAb (green) and Myc-Tag (9B11) Mouse mAb #2276 (red). Overlapping signals are shown in yellow. Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
undefined Image 4: Cas9 and Associated Proteins Antibody Sampler Kit
Confocal immunofluorescent analysis of 293T cells transfected with a construct expressing myc-tagged Cas9 (S. aureus) using Cas9 (S. aureus) (E4G3U) Rabbit mAb (green) and Myc-Tag (9B11) Mouse mAb #2276 (red). Overlapping signals are shown in yellow. Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
undefined Image 5: Cas9 and Associated Proteins Antibody Sampler Kit
Confocal immunofluorescent analysis of Cas9-expressing 293T cells (left) and unmodified 293T cells (right) using Cas9 (S. pyogenes) (D8Y4K) Rabbit mAb (green) and β-Actin (8H10D10) Mouse mAb #3700 (red).
undefined Image 6: Cas9 and Associated Proteins Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

CST Logo
Flow cytometric analysis of 293 cells, untransfected (blue) or transfected with myc-tagged AsCpf1/Cas12a (Strain BV3L6) (green), using AsCpf1/Cas12a (Strain BV3L6) (E1U7C) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
undefined Image 7: Cas9 and Associated Proteins Antibody Sampler Kit
Flow cytometric analysis of 293T cells, untransfected (blue) or transfected with a construct expressing myc-tagged Cas9 (S. aureus) (green), using Cas9 (S. aureus) Rabbit mAb (solid line) compared to a concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed line). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
undefined Image 8: Cas9 and Associated Proteins Antibody Sampler Kit
Flow cytometric analysis of 293T cells, untreated (blue) or transfected with S. pyogenes plasmid (green), using Cas9 (S. pyogenes) (D8Y4K) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
undefined Image 9: Cas9 and Associated Proteins Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

CST Logo
Confocal immunofluorescent analysis of fixed frozen brain from constitutive Cas9 (S. pyogenes)-expressing mice (left, positive) or wild type mice (right, negative), using Cas9 (S. pyogenes) (D8Y4K) Rabbit mAb (green). Samples were mounted in ProLong® Gold Antifade Reagent with DAPI #8961 (blue).
undefined Image 10: Cas9 and Associated Proteins Antibody Sampler Kit
Simple Western™ analysis of lysates (0.001 mg/mL) from COS-7 cells transfected with a construct expressing Cas9 (S. pyogenes) using Cas9 (S. pyogenes) (D8Y4K) Rabbit mAb #65832. The virtual lane view (left) shows a single target band (as indicated) at 1:50 and 1:250 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:50 (blue line) and 1:250 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 66-440 kDa separation module.
undefined Image 11: Cas9 and Associated Proteins Antibody Sampler Kit
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
undefined Image 12: Cas9 and Associated Proteins Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

CST Logo
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing FnCpf1/Cas12a (Strain U112), AsCpf1/Cas12a (Strain BV3L6), Cas9 (S. pyogenes), or Cas9 (S. aureus) (+), using FnCpf1/Cas12a (Strain U112) (E7I2B) Rabbit mAb (upper) and Myc-Tag (71D10) Rabbit mAb #2278 (lower).
undefined Image 13: Cas9 and Associated Proteins Antibody Sampler Kit
Confocal immunofluorescent analysis of 293T cells transfected with a Myc-tagged FnCpf1/Cas12a (Strain U112) construct using FnCpf1/Cas12a (Strain U112) (E7I2B) Rabbit mAb (green) and Myc-Tag (9B11) Mouse mAb #2276 (red). Overlapping signals are shown in yellow. Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
undefined Image 14: Cas9 and Associated Proteins Antibody Sampler Kit
Flow cytometric analysis of 293 cells, untransfected (blue) or transfected with a construct expressing myc-tagged FnCpf1/Cas12a (Strain U112) (green), using FnCpf1/Cas12a (Strain U112) (E7I2B) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
undefined Image 15: Cas9 and Associated Proteins Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.