Overview

  • Product name
    Anti-Cytochrome C antibody [7H8.2C12]
    See all Cytochrome C primary antibodies
  • Description
    Mouse monoclonal [7H8.2C12] to Cytochrome C
  • Host species
    Mouse
  • Tested applications
    Suitable for: Flow Cyt, IHC-Fr, ICC/IF, WB, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Horse, Pigeon, Human, Drosophila melanogaster
  • Immunogen

    Synthetic peptides corresponding to amino acids 1-80, 81-104 and 66-104 of pigeon CYT.

  • Epitope
    The antibody recognizes an epitope within amino acids 93-104 of pigeon Cytochrome C, based on competitive ELISA results.
  • Positive control
    • This antibody gave a positive signal in the following lysates: HeLa whole cell; Jurkat whole cell; Human heart tissue. In Flow Cytometry, this antibody gave a positive signal in methanol fixed/Tween permeabilised HepG2 cells.
  • General notes

    This antibody clone is manufactured by Abcam.

    If you require this antibody in a particular buffer formulation or a particular conjugate for your experiments, please contact orders@abcam.com or you can find further information here.

Properties

Applications

Our Abpromise guarantee covers the use of ab13575 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
Flow Cyt Use 0.1-1µg for 106 cells.

ab170192 - Mouse monoclonal IgG2b, is suitable for use as an isotype control with this antibody.

IHC-Fr Use at an assay dependent concentration.
ICC/IF 1/200 - 1/500. (see Abreviews)
WB Use a concentration of 1 - 5 µg/ml. Detects a band of approximately 15 kDa (predicted molecular weight: 12 kDa).
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Target

  • Function
    Electron carrier protein. The oxidized form of the cytochrome c heme group can accept an electron from the heme group of the cytochrome c1 subunit of cytochrome reductase. Cytochrome c then transfers this electron to the cytochrome oxidase complex, the final protein carrier in the mitochondrial electron-transport chain.
    Plays a role in apoptosis. Suppression of the anti-apoptotic members or activation of the pro-apoptotic members of the Bcl-2 family leads to altered mitochondrial membrane permeability resulting in release of cytochrome c into the cytosol. Binding of cytochrome c to Apaf-1 triggers the activation of caspase-9, which then accelerates apoptosis by activating other caspases.
  • Involvement in disease
    Defects in CYCS are the cause of thrombocytopenia type 4 (THC4) [MIM:612004]; also known as autosomal dominant thrombocytopenia type 4. Thrombocytopenia is the presence of relatively few platelets in blood. THC4 is a non-syndromic form of thrombocytopenia. Clinical manifestations of thrombocytopenia are absent or mild. THC4 may be caused by dysregulated platelet formation.
  • Sequence similarities
    Belongs to the cytochrome c family.
  • Post-translational
    modifications
    Binds 1 heme group per subunit.
  • Cellular localization
    Mitochondrion matrix.
  • Information by UniProt
  • Database links
  • Alternative names
    • CYC antibody
    • CYC_HUMAN antibody
    • CYCS antibody
    • Cytochrome c antibody
    • Cytochrome c somatic antibody
    • HCS antibody
    • THC4 antibody
    see all

Images

  • All lanes : Anti-Cytochrome C antibody [7H8.2C12] (ab13575) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate
    Lane 3 : Human heart tissue lysate - total protein (ab29431)

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Mouse IgG H&L (HRP) preadsorbed (ab97040) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 12 kDa
    Additional bands at: 70 kDa. We are unsure as to the identity of these extra bands.


    Exposure time: 3 minutes


    Abcam recommends using milk (5%) as the blocking agent. Abcam welcomes customer feedback and would appreciate any comments regarding this product and the data presented above.
  • IHC image of Cytochrome C staining in human normal liver formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab13575, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
  • ab13575 staining Cytochrome C in leukocytes from murine bone marrow by Immunocytochemistry/ Immunofluorescence. The cells were fixed in methanol and then blocked using 5% serum for 2 hours at 25°C. Samples were then incubated with primary antibody at 1/250 for 16 hours at 5°C. The secondary antibody used was a goat anti-mouse IgG conjugated to Alexa Fluor® 594 (red) used at a 1/500 dilution. Counterstained with DAPI (blue).

    See Abreview

  • ab13575 staining Cytochrome C from Rat liver by Immunohistochemistry (IHC-Fr-frozen sections).Tissue was fixed with acetone,Samples were incubated with primary antibody (1/300 in PBS 0.1% Triton X) for 15 hours at 4°C.Alexa Fluor® 555 Goat anti-mouse IgG polyclonal (1/1000) was used as the secondary antibody.

     

  • Overlay histogram showing HepG2 cells stained with ab13575 (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 (ab13575, 0.1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1]/mouse IgG2b [PLPV219] (ab91353/ab91366, 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.

  • Ab13575 staining human normal skin tissue. Staining is localised to mitochondria.
    Left panel: with primary antibody at 4 ug/ml. Right panel: isotype control.
    Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 antigen retrieval buffer EDTA pH 9.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.

References

This product has been referenced in:
  • Gopisetty MK  et al. Endoplasmic reticulum stress: major player in size-dependent inhibition of P-glycoprotein by silver nanoparticles in multidrug-resistant breast cancer cells. J Nanobiotechnology 17:9 (2019). Read more (PubMed: 30670028) »
  • Zhang H  et al. Adenovirus-mediated herpes simplex virus thymidine kinase gene therapy combined with ganciclovir induces hepatoma cell apoptosis. Exp Ther Med 17:1649-1655 (2019). Read more (PubMed: 30783433) »
See all 99 Publications for this product

Customer reviews and Q&As

21-30 of 30 Abreviews or Q&A

Application
Western blot
Sample
Mouse Cell lysate - whole cell (liver)
Loading amount
30 µg
Specification
liver
Gel Running Conditions
Reduced Denaturing (4-20% gel)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Dec 13 2010

Application
Western blot
Sample
Mouse Cell lysate - whole cell (brain)
Loading amount
50 µg
Specification
brain
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Aug 25 2010

Application
Immunohistochemistry (Frozen sections)
Sample
Rat Tissue sections (Skeletal Muscle)
Specification
Skeletal Muscle
Fixative
Formaldehyde
Permeabilization
Yes - 0.1% Triton X-100
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Jul 16 2010

Application
Immunocytochemistry/ Immunofluorescence
Sample
Rat Cell (Bone marrow cells)
Specification
Bone marrow cells
Fixative
Formaldehyde
Permeabilization
Yes - 0.1% Triton X-100
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted May 07 2010

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (293 FT)
Specification
293 FT
Fixative
Formaldehyde
Permeabilization
No
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Jan 14 2010

Application
Western blot
Sample
Mouse Tissue lysate - whole (Brain)
Loading amount
10 µg
Specification
Brain
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 24°C

Abcam user community

Verified customer

Submitted Sep 07 2009

Application
Western blot
Sample
Human Cell lysate - other (huh-7)
Loading amount
15 µg
Specification
huh-7
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 20 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C

Abcam user community

Verified customer

Submitted Sep 01 2009

Answer

We also see a 55 kDa band with this antibody (please see the image on the data sheet). Cytochrome c forms aggregates (J aggregates) and  higher molecular weight complexes in the cell and can also aggregate (dimerize, heterodimerize) during the lysis. If the customer were to use  cytochrome c protein they would see also higher molecular weight bands with the antibody.   The customer may need to optimize their lysis to reduce higher molecular weight bands. However, they may not be able to eliminate the higher molecular weight bands totally, there can still be aggregates in the presence of DTT.

Read More
Application
Western blot
Sample
Human Tissue lysate - whole (Brain)
Loading amount
10 µg
Specification
Brain
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 24°C

Abcam user community

Verified customer

Submitted Mar 25 2009

Application
Western blot
Sample
Human Cell lysate - other (hela cells)
Loading amount
20 µg
Specification
hela cells
Treatment
50 MNNG for 20h
Blocking step
BSA as blocking agent for 10 hour(s) and 0 minute(s) · Concentration: 3%

Abcam user community

Verified customer

Submitted Apr 16 2007

21-30 of 30 Abreviews or Q&A

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