Name: Anti-Hexokinase II antibody [EPR20839]
Brand: RabMAb
SKU: ab209847
Rating: 5 (2 reviews)

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Western blot - Anti-Hexokinase II antibody [EPR20839] (ab209847)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EPR20839] to Hexokinase II
Suitable for: WB, IHC-P, ICC/IF, IP, Flow Cyt (Intra)
Knockout validated
Reacts with: Mouse, Rat, Human

Alexa Fluor® 647 Carrier Free HRP

Product name

Anti-Hexokinase II antibody [EPR20839]
See all Hexokinase II primary antibodies
Description

Rabbit monoclonal [EPR20839] to Hexokinase II
Host species

Rabbit
Specificity

Not recommended for rat and mouse tissues in IHC.
Tested applications

Suitable for: WB, IHC-P, ICC/IF, IP, Flow Cyt (Intra)more details
Species reactivity

Reacts with: Mouse, Rat, Human
Immunogen

Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: Hexokinase II recombinant protein (aa16-458); HEK-293, HCT116, HeLa, SH-SY5Y and NIH/3T3 whole cell lysates; HKDC1 and Hexokinase I-knockdown HeLa cell lysates; Rat testis and muscle lysates; Human testis, colon and fetal heart lysates; Mouse muscle lysate. IHC-P: Human heart and pancreatic adenocarcinoma tissues. ICC/IF: HeLa and NIH/3T3 cells. Flow Cyt (intra): HeLa cells. IP: HeLa whole cell lysate.
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
For more information see here.
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

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: PBS, 40% Glycerol, 0.05% BSA
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR20839
Isotype

IgG
Research areas
- Neuroscience
- Development

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab209847 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	(2)	1/1000. Detects a band of approximately 102 kDa (predicted molecular weight: 102 kDa).
IHC-P		1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. Not recommended for rat and mouse tissues.
ICC/IF		1/100.
IP		1/30.
Flow Cyt (Intra)		1/60.

Notes
WB 1/1000. Detects a band of approximately 102 kDa (predicted molecular weight: 102 kDa).
IHC-P 1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. Not recommended for rat and mouse tissues.
ICC/IF 1/100.
IP 1/30.
Flow Cyt (Intra) 1/60.

Tissue specificity

Predominant hexokinase isozyme expressed in insulin-responsive tissues such as skeletal muscle.
Pathway

Carbohydrate metabolism; hexose metabolism.
Sequence similarities

Belongs to the hexokinase family.
Contains 2 hexokinase domains.
Domain

The N- and C-terminal halves of this hexokinase show extensive sequence similarity to each other. The catalytic activity is associated with the C-terminus while regulatory function is associated with the N-terminus. Each domain can bind a single glucose and Gluc-6-P molecule.
Cellular localization

Mitochondrion outer membrane. Its hydrophobic N-terminal sequence may be involved in membrane binding.
Target information above from: UniProt accession P52789 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 3099 Human
- Entrez Gene: 15277 Mouse
- Entrez Gene: 25059 Rat
- Omim: 601125 Human
- SwissProt: P52789 Human
- SwissProt: O08528 Mouse
- SwissProt: P27881 Rat
- Unigene: 406266 Human
- Unigene: 591588 Human
- Unigene: 255848 Mouse
- Unigene: 469167 Mouse
- Unigene: 91375 Rat
see all
Alternative names
- DKFZp686M1669 antibody
- Hexokinase 2 antibody
- Hexokinase 2 muscle antibody
- Hexokinase type II antibody
- Hexokinase-2 antibody
- HK 2 antibody
- HK II antibody
- HK2 antibody
- HKII antibody
- HxK 2 antibody
- HxK2 antibody
- HXK2_HUMAN antibody
- Muscle form hexokinase antibody
see all

Western blot - Anti-Hexokinase II antibody [EPR20839] (ab209847)

All lanes : Anti-Hexokinase II antibody [EPR20839] (ab209847) at 1/1000 dilution

Lane 1 : Wild-type HCT116 cell lysate
Lane 2 : HK2 knockout HCT116 cell lysate
Lane 3 : HeLa cell lysate
Lane 4 : SH-SY5Y cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 102 kDa
Observed band size: 105 kDa why is the actual band size different from the predicted?

Lanes 1 - 4: Merged signal (red and green). Green - ab209847 observed at 105 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab209847 was shown to react with Anti-Hexokinase II in HCT 116 wild-type cells in western blot with loss of signal observed in HK2 knockout cell line ab273721 (HK2 knockout cell lysate ab275250). HCT 116 wild-type and HK2 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab209847 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
Immunocytochemistry/ Immunofluorescence - Anti-Hexokinase II antibody [EPR20839] (ab209847)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cells labeling Hexokinase II with ab209847 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Cytoplasmic staining in HeLa cells.

The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hexokinase II antibody [EPR20839] (ab209847)

Immunohistochemical analysis of paraffin-embedded human pancreatic adenocarcinoma tissue labeling Hexokinase II with ab209847 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Strong cytoplasmic staining in human pancreatic adenocarcinoma (PMID: 26137268) 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) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Flow Cytometry (Intracellular) - Anti-Hexokinase II antibody [EPR20839] (ab209847)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cell line labeling Hexokinase II with ab209847 at 1/60 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody.
Western blot - Anti-Hexokinase II antibody [EPR20839] (ab209847)

All lanes : Anti-Hexokinase II antibody [EPR20839] (ab209847) at 1/1000 dilution

Lane 1 : Wild-type HEK-293 cell lysate
Lane 2 : HK2 knockout HEK-293 cell lysate
Lane 3 : HeLa cell lysate
Lane 4 : A549 cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 102 kDa
Observed band size: 110 kDa why is the actual band size different from the predicted?

Lanes 1 - 4: Merged signal (red and green). Green - ab209847 observed at 110 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab209847 was shown to react with Hexokinase II in wild-type HEK-293 cells in western blot with loss of signal observed in HK2 knockout sample. Wild-type and HK2 knockout HEK-293 cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab209847 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 1000 Dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
Western blot - Anti-Hexokinase II antibody [EPR20839] (ab209847)

All lanes : Anti-Hexokinase II antibody [EPR20839] (ab209847) at 1/1000 dilution

Lane 1 : HeLa (human epithelial cell line from cervix adenocarcinoma), whole cell lysate
Lane 2 : HKDC1-knockdown HeLa (human epithelial cell line from cervix adenocarcinoma) cell lysate
Lane 3 : Hexokinase I-knockdown HeLa (human epithelial cell line from cervix adenocarcinoma) cell lysate
Lane 4 : Hexokinase II-knockdown HeLa (human epithelial cell line from cervix adenocarcinoma) cell lysate

Lysates/proteins at 10 µg per lane.

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

Developed using the ECL technique.

Predicted band size: 102 kDa
Observed band size: 102 kDa

Exposure time: 41 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

The blot was developed on a BIO-RAD^® ChemiDoc™ MP instrument.
Immunoprecipitation - Anti-Hexokinase II antibody [EPR20839] (ab209847)

Hexokinase II was immunoprecipitated from 0.35 mg HeLa (humanepithelial cell line from cervix adenocarcinoma) whole cell lysate with ab209847 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab209847 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1: HeLa whole cell lysate 10 μg (Input).
Lane 2: ab209847 IP in HeLa whole cell lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab209847 in HeLa whole cell lysate.

Exposure time: 30 seconds.

Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Immunocytochemistry/ Immunofluorescence - Anti-Hexokinase II antibody [EPR20839] (ab209847)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embryonic fibroblast cell line) cells labeling Hexokinase II with ab209847 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Cytoplasmic staining in NIH/3T3 cells.

The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.
Western blot - Anti-Hexokinase II antibody [EPR20839] (ab209847)

All lanes : Anti-Hexokinase II antibody [EPR20839] (ab209847) at 1/1000 dilution

Lane 1 : Rat testis lysate at 20 µg
Lane 2 : Rat muscle lysate at 20 µg
Lane 3 : SH-SY5Y (human neuroblastoma cell line from bone marrow), whole cell lysate at 20 µg
Lane 4 : Human testis lysate at 20 µg
Lane 5 : Mouse muscle lysate at 20 µg
Lane 6 : Human colon lysate at 10 µg
Lane 7 : Human fetal heart lysate at 10 µg
Lane 8 : NIH/3T3 (mouse embryonic fibroblast cell line), whole cell lysate at 10 µg

Secondary
Lanes 1-5 & 8 : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Lanes 6-7 : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/4000 dilution

Developed using the ECL technique.

Predicted band size: 102 kDa
Observed band size: 102 kDa

Exposure times: Lanes 1-3,6,7: 3 minutes; Lanes 4,5,8: 58 seconds.

Blocking/Dilution buffer: 5% NFDM/TBST.

The blot was developed on a BIO-RAD^® ChemiDoc™ MP instrument.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Hexokinase II antibody [EPR20839] (ab209847)

Immunohistochemical analysis of paraffin-embedded human heart tissue labeling Hexokinase II with ab209847 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Granularly cytoplasmic staining in human heart (PMID: 4058069; PMID: 26722360) 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) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-Hexokinase II antibody [EPR20839] (ab209847)

All lanes : Anti-Hexokinase II antibody [EPR20839] (ab209847) at 1/10000 dilution

Lane 1 : HKDC1 recombinant protein (aa16-458)
Lane 2 : Hexokinase II recombinant protein (aa16-458)
Lane 3 : Hexokinase I recombinant protein (aa16-458)

Lysates/proteins at 0.01 µg per lane.

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

Developed using the ECL technique.

Predicted band size: 102 kDa
Observed band size: 50 kDa why is the actual band size different from the predicted?

Exposure time: 3 minutes

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

The blot was developed on a BIO-RAD^® ChemiDoc™ MP instrument.
Anti-Hexokinase II antibody [EPR20839] (ab209847)

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

SDS download

Country/Region

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Datasheet download

Download

Publishing research using ab209847? Please let us know so that we can cite the reference in this datasheet.

ab209847 has been referenced in 49 publications.

Sun J et al. aFGF alleviates diabetic endothelial dysfunction by decreasing oxidative stress via Wnt/ß-catenin-mediated upregulation of HXK2. Redox Biol 39:101811 (2021). PubMed: 33360774
Li Y et al. Knockdown of circPUM1 impedes cell growth, metastasis and glycolysis of papillary thyroid cancer via enhancing MAPK1 expression by serving as the sponge of miR-21-5p. Genes Genomics 43:141-150 (2021). PubMed: 33481227
Li X et al. NR2F1-AS1/miR-140/HK2 Axis Regulates Hypoxia-Induced Glycolysis and Migration in Hepatocellular Carcinoma. Cancer Manag Res 13:427-437 (2021). PubMed: 33488124
Wang F et al. A positive feedback loop between TAZ and miR-942-3p modulates proliferation, angiogenesis, epithelial-mesenchymal transition process, glycometabolism and ROS homeostasis in human bladder cancer. J Exp Clin Cancer Res 40:44 (2021). PubMed: 33499877
Liu J et al. Circ_0105346 Knockdown Inhibits Osteosarcoma Development via Regulating miR-1182/WNT7B Axis. Cancer Manag Res 13:521-535 (2021). PubMed: 33505171

View all Publications for this product

Customer reviews and Q&As

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1-2 of 2 Abreviews

Application

Western blot

Sample

Mouse Tissue lysate - whole (brain)

Gel Running Conditions

Reduced Denaturing (TEO TRICINE 4-20% gel (ab119209))

Loading amount

20 µg

Specification

brain

Blocking step

BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Dr. Yonatan Feuermann

Verified customer

Submitted Apr 12 2021

Application

Western blot

Sample

Human Cell lysate - whole cell (Induced pluripotent stem cells (iPSCs))

Gel Running Conditions

Reduced Denaturing (10%)

Loading amount

25 µg

Specification

Induced pluripotent stem cells (iPSCs)

Blocking step

Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 23°C

The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

HERR DR. Vladimir Milenkovic

Verified customer

Submitted Jun 30 2020

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Overview

Product name

Description

Host species

Specificity

Tested applications

Species reactivity

Immunogen

Positive control

General notes

Properties

Form

Storage instructions

Storage buffer

Purity

Clonality

Clone number

Isotype

Research areas

Associated products

Alternative Versions

Compatible Secondaries

Isotype control

KO cell lines

KO cell lysates

Recombinant Protein

Related Products

Applications

The Abpromise guarantee

Target

Tissue specificity

Pathway

Sequence similarities

Domain

Cellular localization

Database links

Alternative names

Images

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Datasheets and documents

SDS download

Datasheet download

Certificate of Compliance

References (49)

Customer reviews and Q&As

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Western blot abreview for Anti-Hexokinase II antibody [EPR20839]

Western blot abreview for Anti-Hexokinase II antibody [EPR20839]