Overview

  • Product name
    Anti-ROCK1 antibody [EP786Y]
    See all ROCK1 primary antibodies
  • Description
    Rabbit monoclonal [EP786Y] to ROCK1
  • Host species
    Rabbit
  • Specificity
    This antibody recognizes both the cleaved C-terminus of ROCK 1 (30 kDa) and full length protein (158 kDa). The immunogen used for this product shares 83% homology with ROCK2. Cross-reactivity with this protein has not been confirmed experimentally.
  • Tested applications
    Suitable for: IHC-Fr, WB, IHC-P, ICC/IF, IP, Flow Cytmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Cow, Human
  • Immunogen

    Synthetic peptide within Human ROCK1 aa 1100-1200 (C terminal). The exact sequence is proprietary.
    Database link: Q13464

  • Positive control
    • WB: Untreated, Calyculin A treated and Camptothecin treated HeLa whole cell lysate (ab150035). Jurkat, Ramos, PC-12 and RAW264.7 cell lysates. IHC-P: Human adenocarcinoma of the colon and thyroid gland carcinoma tissues. ICC/IF: HeLa cells. Flow Cyt: HeLa cells. IP: HeLa whole cell lysate (ab150035).
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab45171 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
IHC-Fr 1/100. PubMed: 19295659
WB 1/2000 - 1/10000. Predicted molecular weight: 158 kDa.

For unpurified use at 1/500.

IHC-P 1/50 - 1/250. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

ICC/IF 1/50 - 1/500.

 

IP 1/40 - 1/50.

 

Flow Cyt 1/1000.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

Target

  • Function
    Protein kinase which is a key regulator of actin cytoskeleton and cell polarity. Involved in regulation of smooth muscle contraction, actin cytoskeleton organization, stress fiber and focal adhesion formation, neurite retraction, cell adhesion and motility via phosphorylation of DAPK3, GFAP, LIMK1, LIMK2, MYL9/MLC2, PFN1 and PPP1R12A. Phosphorylates FHOD1 and acts synergistically with it to promote SRC-dependent non-apoptotic plasma membrane blebbing. Phosphorylates JIP3 and regulates the recruitment of JNK to JIP3 upon UVB-induced stress. Acts as a suppressor of inflammatory cell migration by regulating PTEN phosphorylation and stability. Acts as a negative regulator of VEGF-induced angiogenic endothelial cell activation. Required for centrosome positioning and centrosome-dependent exit from mitosis. Plays a role in terminal erythroid differentiation. May regulate closure of the eyelids and ventral body wall by inducing the assembly of actomyosin bundles. Promotes keratinocyte terminal differentiation. Involved in osteoblast compaction through the fibronectin fibrillogenesis cell-mediated matrix assembly process, essential for osteoblast mineralization.
  • Tissue specificity
    Detected in blood platelets.
  • Sequence similarities
    Belongs to the protein kinase superfamily. AGC Ser/Thr protein kinase family.
    Contains 1 AGC-kinase C-terminal domain.
    Contains 1 PH domain.
    Contains 1 phorbol-ester/DAG-type zinc finger.
    Contains 1 protein kinase domain.
    Contains 1 REM (Hr1) repeat.
  • Domain
    The C-terminal auto-inhibitory domain interferes with kinase activity. RHOA binding leads to a conformation change and activation of the kinase. Truncated ROCK1 is constitutively activated.
  • Post-translational
    modifications
    Autophosphorylated on serine and threonine residues.
    Cleaved by caspase-3 during apoptosis. This leads to constitutive activation of the kinase and membrane blebbing.
  • Cellular localization
    Cytoplasm. Cytoplasm, cytoskeleton, microtubule organizing center, centrosome, centriole. Golgi apparatus membrane. Cell projection, bleb. Cytoplasm, cytoskeleton. Cell membrane. Cell projection, lamellipodium. Cell projection, ruffle. Associated with the mother centriole and an intercentriolar linker. Colocalizes with ITGB1BP1 and ITGB1 at the cell membrane predominantly in lamellipodia and membrane ruffles, but also in retraction fibers. Localizes at the cell membrane in an ITGB1BP1-dependent manner (By similarity). A small proportion is associated with Golgi membranes.
  • Information by UniProt
  • Database links
  • Alternative names
    • coiled-coil-containing protein kinase 1 antibody
    • coiled-coil-containing protein kinase I antibody
    • MGC131603 antibody
    • MGC43611 antibody
    • p160 Rhoassociated coiled coil-forming protein kinase antibody
    • p160 ROCK-1 antibody
    • p160 ROCK1 antibody
    • p160ROCK antibody
    • PRO0435 antibody
    • Renal carcinoma antigen NY REN 35 antibody
    • Renal carcinoma antigen NY-REN-35 antibody
    • Rho associated coiled coil containing protein kinase 1 antibody
    • Rho associated protein kinase 1 antibody
    • Rho kinase antibody
    • Rho-alpha kinase antibody
    • Rho-associated antibody
    • Rho-associated protein kinase 1 antibody
    • ROCK I antibody
    • ROCK-I antibody
    • ROCK1 antibody
    • ROCK1_HUMAN antibody
    • Rok antibody
    • rokalpha antibody
    see all

Images

  • All lanes : Anti-ROCK1 antibody [EP786Y] (ab45171) at 1/5000 dilution (purified)

    Lane 1 : HeLa cell lysate - treated with Calyculin A
    Lane 2 : HeLa cell lysate - treated with Camptothecin
    Lane 3 : HeLa cell lysate
    Lane 4 : Jurkat cell lysate
    Lane 5 : Ramos cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Peroxidase-conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

    Predicted band size: 158 kDa
    Observed band size: 158 kDa



    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human adenocarcinoma of the colon tissue labelling ROCK1 with purified ab45171 at 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. A prediluted HRP-polymer conjugated anti-rabbit IgG was used as the secondary antibody. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling ROCK1 with purified ab45171 at 1/50. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150078, an Alexa Fluor® 555-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain.

    Control: primary antibody (1/50) and secondary antibody, ab150113, an Alexa Fluor® 488-conjugated goat anti-mouse IgG (1/500).

  • Overlay histogram showing HeLa cells stained with unpurified ab45171 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab45171, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

  • ab45171 (purified) at 1/40 immunoprecipitating ROCK1 in HeLa cell lysate. For western blotting, a peroxidase-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/1000).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • All lanes : Anti-ROCK1 antibody [EP786Y] (ab45171) at 1/5000 dilution (purified)

    Lane 1 : PC-12 cell lysate
    Lane 2 : RAW264.7 cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Peroxidase-conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

    Predicted band size: 158 kDa
    Observed band size: 158 kDa



    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • Unpurified ab45171 staining ROCK1 in mouse embyro 18dpc tissue sections by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% FBS/BSA for 2 hours at room temperature; antigen retrieval was by heat mediation in Tris pH 9. Samples were incubated with primary antibody (1/100 in 1% BSA + 1% FBS in TBS) for 16 hours. An undiluted HRP-conjugated goat anti-rabbit IgG polyclonal was used as the secondary antibody.

    See Abreview

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human thyroid gland carcinoma tissue labelling ROCK1 with unpurified ab45171.

References

This product has been referenced in:
  • Roberto GM  et al. MiR-708-5p is inversely associated with EWS/FLI1 Ewing sarcoma but does not represent a prognostic predictor. Cancer Genet 230:21-27 (2019). Read more (PubMed: 30470587) »
  • Yang Z  et al. DLC2 operates as a tumor suppressor gene in breast cancer via the RhoGTPase pathway. Oncol Lett 17:2107-2116 (2019). Read more (PubMed: 30719106) »
See all 41 Publications for this product

Customer reviews and Q&As

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1-6 of 6 Abreviews

Application
Western blot
Sample
Pig Cell lysate - whole cell (heart)
Gel Running Conditions
Reduced Denaturing
Loading amount
5 µg
Specification
heart
Blocking step
Licor as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 24°C

Abcam user community

Verified customer

Submitted Nov 04 2016

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step
FBS / BSA as blocking agent for 3 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: RT°C
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Tris pH 9
Sample
Human Tissue sections (Infantile fibromatosis)
Specification
Infantile fibromatosis
Permeabilization
No
Fixative
Formaldehyde

Abcam user community

Verified customer

Submitted Jul 04 2014

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step
FBS / BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: RT°C
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Tris pH 9
Sample
Mouse Tissue sections (Mouse embryo 18dpc)
Specification
Mouse embryo 18dpc
Permeabilization
No
Fixative
Formaldehyde

Abcam user community

Verified customer

Submitted Jan 15 2014

Application
Western blot
Sample
Mouse Cell lysate - whole cell (MEF)
Loading amount
30 µg
Specification
MEF
Treatment
TGF b1 o/n 5 ug/ul
Gel Running Conditions
Reduced Denaturing (7.5)
Blocking step
(agent) for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Feb 27 2013

Application
Immunocytochemistry/ Immunofluorescence
Sample
Rhesus monkey Cell (Primary Rhesus macaque Intestinal Macrophages)
Specification
Primary Rhesus macaque Intestinal Macrophages
Fixative
Paraformaldehyde
Permeabilization
Yes - 0.1% Triton-X-100
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Aug 16 2011

Application
Western blot
Sample
Human Recombinant protein (vero cells expressing ROCK1 on pcDNA vector)
Loading amount
20 µg
Specification
vero cells expressing ROCK1 on pcDNA vector
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Dec 17 2009

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