Overview

  • Product name
    Anti-Ki67 antibody [SP6]
    See all Ki67 primary antibodies
  • Description
    Rabbit monoclonal [SP6] to Ki67
  • Tested applications
    Suitable for: IHC-FoFr, ICC/IF, Flow Cyt, IHC-Fr, WB, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human, Common marmoset
  • Immunogen

    Synthetic peptide within Human Ki67 aa 2300-2400 (C terminal). The exact sequence is proprietary.
    Database link: P46013

  • Epitope
    C-terminus
  • Positive control
    • IHC-P: Human tonsil and testis tissue. Common marmoset spleen tissue. Rat esophagus, small intestine and liver tissue. Mouse embryonic skin tissue. Transgenic mouse spinal cord tissue. ICC/IF: HAP1 cells. Human cardiac stem cells. HEp-2 cells. Flow Cyt: HAP1 cells.

Properties

Applications

Our Abpromise guarantee covers the use of ab16667 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-FoFr 1/5000.

Antigen retrieval: Boil tissue section in 10mM citrate buffer, pH 6.0 for 10 min followed by cooling at room temperature for 20 min.

ICC/IF 1/250.
Flow Cyt Use a concentration of 1 µg/ml.

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

IHC-Fr 1/1000. A higher dilution is recommended for frozen tissues than for FFPE tissues. We suggest a starting dilution of 1/500.
WB Use at an assay dependent concentration. PubMed: 20562294
IHC-P 1/100.

Target

  • Function
    Required to maintain individual mitotic chromosomes dispersed in the cytoplasm following nuclear envelope disassembly (PubMed:27362226). Associates with the surface of the mitotic chromosome, the perichromosomal layer, and covers a substantial fraction of the chromosome surface (PubMed:27362226). Prevents chromosomes from collapsing into a single chromatin mass by forming a steric and electrostatic charge barrier: the protein has a high net electrical charge and acts as a surfactant, dispersing chromosomes and enabling independent chromosome motility (PubMed:27362226). Binds DNA, with a preference for supercoiled DNA and AT-rich DNA (PubMed:10878551). Does not contribute to the internal structure of mitotic chromosomes (By similarity). May play a role in chromatin organization (PubMed:24867636). It is however unclear whether it plays a direct role in chromatin organization or whether it is an indirect consequence of its function in maintaining mitotic chromosomes dispersed.
  • Sequence similarities
    Contains 1 FHA domain.
    Contains 16 K167R repeats.
    Contains 1 PP1-binding domain.
  • Developmental stage
    Expression occurs preferentially during late G1, S, G2 and M phases of the cell cycle, while in cells in G0 phase the antigen cannot be detected (at protein level) (PubMed:6206131). Present at highest level in G2 phase and during mitosis (at protein level). In interphase, forms fiber-like structures in fibrillarin-deficient regions surrounding nucleoli (PubMed:2674163, PubMed:8799815).
  • Post-translational
    modifications
    Phosphorylated. Hyperphosphorylated in mitosis (PubMed:10502411, PubMed:10653604). Hyperphosphorylated form does not bind DNA.
  • Cellular localization
    Chromosome. Nucleus. Nucleus, nucleolus. Associates with the surface of the mitotic chromosome, the perichromosomal layer, and covers a substantial fraction of the mitotic chromosome surface (PubMed:27362226). Associates with satellite DNA in G1 phase (PubMed:9510506). Binds tightly to chromatin in interphase, chromatin-binding decreases in mitosis when it associates with the surface of the condensed chromosomes (PubMed:15896774, PubMed:22002106). Predominantly localized in the G1 phase in the perinucleolar region, in the later phases it is also detected throughout the nuclear interior, being predominantly localized in the nuclear matrix (PubMed:22002106).
  • Information by UniProt
  • Database links
  • Alternative names
    • Antigen identified by monoclonal antibody Ki 67 antibody
    • Antigen identified by monoclonal antibody Ki-67 antibody
    • Antigen KI-67 antibody
    • Antigen KI67 antibody
    • Antigen Ki67 antibody
    • KI67_HUMAN antibody
    • KIA antibody
    • Marker of proliferation Ki-67 antibody
    • MIB 1 antibody
    • MIB antibody
    • MKI67 antibody
    • PPP1R105 antibody
    • Proliferation marker protein Ki-67 antibody
    • Proliferation related Ki 67 antigen antibody
    • Protein phosphatase 1 regulatory subunit 105 antibody
    • RP11-380J17.2 antibody
    see all

Images

  • Overlay histogram showing HAP1 wildtype (green line) and HAP1-MKI67 knockout cells (red line) stained with ab16667. The cells were fixed with  80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab16667,1µg/ml) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150081) at 1/2000 dilution for 30 min at 22°C.
    A Rabbit IgG isotype control antibody  (ab172730) was used at the same concentration and conditions as the primary antibody (HAP1 wildtype - black line, HAP1-MKI67 knockout - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).
    Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.
  • Immunohistochemical analysis of human tonsil tissue labeling Ki-67 with ab16667 at 1/200. The HRP/AEC-staining procedure was used for detection.

  • ab16667 staining Ki67 in wild-type HAP1 cells (top panel) and Ki67 knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol for 5 minutes, permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1 hour. The cells were then incubated with ab16667 at 1/250 dilution and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1 hour with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

  • ab16667 staining Ki67 in common marmoset spleen by immunohistochemistry (formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer. Samples were then blocked with 1% BSA for 10 minutes at 21ºC followed by incubation with the primary antibody for 2 hours at 1/100. A biotin-conjugated goat anti-rabbit polyclonal was used as secondary antibody at a 1/250 dilution.

    See Abreview

  • Immunocytochemistry/ Immunofluorescence analysis of human cardiac stem cells labeling Ki67 with ab16667 at 1/250 dilution. Cells were fixed in paraformaldehyde and permeabilized with Triton x-100, 0.01%. Cells were blocked in BSA for 1 hour at room temperature. A polyclonal chicken anti-rabbit Alex Fluor® 488 secondary antibody was used at 1/500 dilution. 

    See Abreview

  • ab16667 staining Ki67 in rat liver tissue sections by immunohistochemistry (IHC-P - formaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA for 2 hours at 22°C. Samples were incubated with primary antibody (1/500 in PBS-T + 1% BSA) for 18 hours at 4°C. A biotin-conjugated goat anti-rabbit IgG monoclonal (1/2000) was used as the secondary antibody.

    See Abreview

  • ab16667 staining Ki67 in human testis by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer. Samples were then blocked with 1% BSA for 10 minutes at 21ºC followed by incubation with the primary antibody for 2 hours at 1/100. A biotin-conjugated goat anti-rabbit polyclonal was used as secondary antibody at a 1/250 dilution.

    See Abreview

  • ab16667 staining Ki67 in mouse embryonic skin tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer. Samples were then blocked with 1% BSA for 10 minutes at 21ºC followed by incubation with the primary antibody for 16 hours at 1/50. A biotin-conjugated goat anti-rabbit polyclonal was used as secondary antibody at a 1/250 dilution.

    See Abreview

  • ab16667 staining Ki67 in rat oesophagus by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer. Samples were then blocked with 1% BSA for 10 minutes at 21ºC followed by incubation with the primary antibody for 30 minutes at 1/100. A biotin-conjugated goat anti-rabbit polyclonal was used as secondary antibody at a 1/250 dilution.

    See Abreview

  • ab16667 staining Ki67 in rat small intestine tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using 10 mM citrate buffer pH 6.0. Samples were then blocked with 10% serum for 20 minutes at room temperature followed by incubation with the undiluted primary antibody for 30 minutes. A biotin-conjugated goat anti-rabbit polyclonal was used as secondary antibody at a 1/2000 dilution.

    See Abreview

  • ab16667 staining Ki67 - Proliferation Marker in human HEp-2 cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde and permeabilized with Triton X-100 0.25% in PBS. Samples were incubated with primary antibody (1/50 in DPBS) for 1 hour at 21°C. An Atto488-conjugated Donkey anti-rabbit polyclonal (1/50) was used as the secondary antibody.

    See Abreview

  • ab16667 staining Ki67 (red) in transgenic mouse spinal cord tissue sections (depleted of oligodendrocytes) by Immunohistochemistry (PFA perfusion fixed frozen sections). Tissue samples were fixed by perfusion with paraformaldehyde, permeablized with 0.5% Triton X-100 and blocked with 10% serum for 1 hour at 25°C; antigen retrieval was by heat mediation in 10 mM citrate buffer, pH 6, for 20 minutes at 97°C in a water bath. The sample was incubated with primary antibody (1/300 in PBS + 0.1% Triton X-100 + 1% serum) at 25°C for 16 hours. An Alexa Fluor® 594-conjugated donkey anti-rabbit IgG (H+L) polyclonal (1/700) was used as the secondary antibody. Counterstained with Iba1 (green) a marker for microglia and DAPI.

    See Abreview

References

This product has been referenced in:
  • Fernández-Gil B  et al. Melatonin protects rats from radiotherapy-induced small intestine toxicity. PLoS One 12:e0174474 (2017). IHC-P ; Rat . Read more (PubMed: 28403142) »
  • Liu Q  et al. Plasmodium parasite as an effective hepatocellular carcinoma antigen glypican-3 delivery vector. Oncotarget 8:24785-24796 (2017). IHC . Read more (PubMed: 28445973) »

See all 371 Publications for this product

Customer reviews and Q&As

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Skin)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: ph6 citrate
Permeabilization
No
Specification
Skin
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: 25°C
Fixative
Formaldehyde
Username

Dr. Victoria Thompson

Verified customer

Submitted Jun 29 2016

Either ab16667 or ab27619 would work for your chosen application and species.

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Colon)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: pH 6.0 Citrate
Permeabilization
No
Specification
Colon
Blocking step
No blocking step used for 10 minute(s) · Concentration: 1% · Temperature: RT°C
Fixative
Formaldehyde
Username

Abcam user community

Verified customer

Submitted Nov 14 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (lung carcinoma)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Tris/EDTA pH 9.0
Permeabilization
No
Specification
lung carcinoma
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 15% · Temperature: RT°C
Fixative
10% NBF
Username

Abcam user community

Verified customer

Submitted Aug 21 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (spleen)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Tris/EDTA pH 9.0
Permeabilization
No
Specification
spleen
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 15% · Temperature: RT°C
Fixative
10% NBF
Username

Abcam user community

Verified customer

Submitted Aug 19 2017

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Pig Tissue sections (Liver)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citrate buffer (pH 6.0)
Permeabilization
No
Specification
Liver
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: 22°C
Fixative
Formaldehyde
Username

Abcam user community

Verified customer

Submitted Aug 08 2017

Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Molar)
Permeabilization
Yes - Triton X-100
Specification
Molar
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: RT°C
Fixative
Paraformaldehyde
Username

Dr. Tina Yuan

Verified customer

Submitted Apr 25 2017

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (Cell line)
Permeabilization
Yes - 0.5% Titon
Specification
Cell line
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative
Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted Apr 17 2017

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HEK293)
Permeabilization
Yes - 0.1% Triton X100
Specification
HEK293
Blocking step
Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative
Formaldehyde
Username

Abcam user community

Verified customer

Submitted Apr 10 2017

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Cow Tissue sections (ovary)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: 10 mM citrate, pH6.0
Permeabilization
No
Specification
ovary
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: 25°C
Fixative
Formaldehyde
Username

Abcam user community

Verified customer

Submitted Feb 09 2017

1-10 of 100 Abreviews or Q&A

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