Revision 3

#51509

Store at -20C

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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.

Applications:
W, IP

Reactivity:
H M R Mk

Sensitivity:
Endogenous

MW (kDa):
175

Source/Isotype:
Rabbit IgG

UniProt ID:
#O14976

Entrez-Gene Id:
2580

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:50

Storage

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 antibody.

Specificity/Sensitivity

GAK (E7W7Z) Rabbit Monoclonal Antibody recognizes endogenous levels of total GAK protein. A band of unknown origin is detected at ~45 kDa and at ~20kDa.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro758 of human GAK protein.

Background

Cyclin-G-associated kinase (GAK) is the ubiquitous form of the neuronal-specific protein auxilin 1 (1) and is an essential cofactor for Hsc70-dependent uncoating of clathrin-coated vesicles in non-neuronal cells (2). The recruitment of GAK to clathrin-coated pits during endocytosis is dependent on its PTEN-like domain, which has been shown to bind to phospholipids (3). It has been postulated that the JAK inhibitor baricitinib may impair entry of the SARS-CoV-2 virus by inhibiting AAK1- and GAK-mediated endocytosis via ACE2 (4). In the nucleus, GAK forms complexes with cyclin G1 and other nuclear proteins (5) and plays an essential role in proper mitotic progression by maintaining centrosome structure (6). GAK is also required for the regulation of basal, PRKN-independent mitophagy by modifying the mitochondrial network and lysosomal morphology that compromise efficient transport of mitochondria for degradation (7,8).

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

W: Western Blotting IP: Immunoprecipitation

Cross-Reactivity Key

H: Human M: Mouse R: Rat Mk: Monkey

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.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

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 3

Cell Signaling Technology Logo
Western blot analysis of extracts from various cell lines using GAK (E7W7Z) Rabbit mAb (upper) or GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).
Western Blotting Image 1: GAK (E7W7Z) Rabbit Monoclonal Antibody
Western blot analysis of extracts from NCI-H520 cells, transfected with control siRNA (-) or GAK siRNA (+), using GAK (E7W7Z) Rabbit mAb (upper) or GAPDH (D16H11) XP® Rabbit mAb #5174 (lower). The loss of GAK signal in extracts from GAK siRNA-transfected cells confirms specificity of the antibody for GAK.
Western Blotting Image 2: GAK (E7W7Z) Rabbit Monoclonal Antibody
Western blot analysis of extracts from 293T cells, mock transfected (lane 1) or transiently transfected with a plasmid encoding Myc/DDK-tagged GAK protein (lane 2), using GAK (E7W7Z) Rabbit mAb (upper), Myc-Tag (71D10) Rabbit mAb #2278 (middle), or GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).
Western Blotting Image 3: GAK (E7W7Z) Rabbit Monoclonal Antibody
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 3

Cell Signaling Technology Logo
Immunoprecipitation of GAK protein from KATO III cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is GAK (E7W7Z) Rabbit mAb. Western blot analysis was performed using GAK (E7W7Z) Rabbit mAb. Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb (HRP Conjugate) #5127 was used as a secondary antibody.
Immunoprecipitation Image 1: GAK (E7W7Z) Rabbit Monoclonal Antibody
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.