Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-Ki67 antibody [SP6] - BSA free (ab231172)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [SP6] to Ki67 - BSA free
  • Suitable for: mIHC, Flow Cyt, ICC, IHC-P, WB
  • Knockout validated
  • Reacts with: Mouse, Human, Common marmoset

Overview

  • Product name

    Anti-Ki67 antibody [SP6] - BSA free
    See all Ki67 primary antibodies

  • Description

    Rabbit monoclonal [SP6] to Ki67 - BSA free

  • Host species

    Rabbit

  • Tested Applications & Species

    Application Species
    Flow Cyt
    Human
    ICC
    Human
    IHC-P
    Human
    mIHC
    Human
    WB
    Human
    See all applications and species data

  • Immunogen

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • Epitope

    C-terminus

  • Positive control

    • WB: Ramos and HeLa cell lysates. 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.

  • General notes

    FOR RESEARCH USE ONLY. For commercial use, please contact partnerships@abcam.com.

    ab231172 is the carrier-free version of ab16667. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with

    Ab231172 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    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.

Properties

Images

  • Multiplex immunohistochemistry - Anti-Ki67 antibody [SP6] - BSA free (ab231172)
    Multiplex immunohistochemistry - Anti-Ki67 antibody [SP6] - BSA free (ab231172)

    This data was developed using the same antibody clone in a different buffer formulation (ab16667).

    Fluorescence multiplex immunohistochemical analysis of normal human tonsil tissue (formalin-fixed paraffin-embedded section).

    Merged staining of anti-PD1 (ab237728; orange; Opal™520), anti-PDL1 (ab237726; green; Opal™540), anti-CD68 (ab192847; yellow; Opal™570), anti-CD3 (ab16669; red; Opal™620), anti-Ki67 (ab16667; light blue; Opal™650) and anti-PanCK (ab7753; grey; Opal™690).

    The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 7-color automation IHC kit (NEL821001KT, Akoya Biosciences®).

    The section was incubated in six rounds of staining; in the order of ab237728 (1/500 dilution), ab237726 (1/500 dilution), ab192847 (1/300 dilution), ab16669 (1/300 dilution), ab16667 (1/200 dilution) and ab7753 (1/200 dilution); each using a separate fluorescent tyramide signal amplification system.

    Sodium citrate antigen retrieval (Leica ER1, pH6.0, 30 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity.

    DAPI (dark blue) was used as a nuclear counter stain.

    Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra Polaris.

    This image was generated from the hybridoma version.

  • Western blot - Anti-Ki67 antibody [SP6] - BSA free (ab231172)
    Western blot - Anti-Ki67 antibody [SP6] - BSA free (ab231172)
    All lanes : Anti-Ki67 antibody [SP6] (ab16667) at 1/100 dilution

    Lane 1 : Ramos cell lysate
    Lane 2 : Wild-type HeLa cell lysate
    Lane 3 : Ki67 knockout HeLa cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 358 kDa
    Observed band size: 359 kDa
    why is the actual band size different from the predicted?



    This data was developed using the same antibody clone in a different buffer formulation (ab16667).

    Lanes 1-3: Merged signal (red and green). Green - ab16667 observed at 359 kDa. Red - loading control, ab130007 observed at 125 kDa.

    ab16667 was shown to react with Ki67 in wild-type HeLa. Loss of signal was observed when knockout sample ab263762 was used. Wild-type and Ki67 knockout samples were subjected to SDS-PAGE. ab16667 and Anti-Vinculin antibody VIN-54] (ab130007) were incubated overnight at 4°C at 1 in 100 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

  • Immunocytochemistry - Anti-Ki67 antibody [SP6] - BSA free (ab231172)
    Immunocytochemistry - Anti-Ki67 antibody [SP6] - BSA free (ab231172)

    This data was developed using the same antibody clone in a different buffer formulation (ab16667).

     

    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 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) secondary antibody at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

     

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Ki67 antibody [SP6] - BSA free (ab231172)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Ki67 antibody [SP6] - BSA free (ab231172)

    This data was developed using the same antibody clone in a different buffer formulation (ab16667).

    IHC image of ab16667 staining Ki67 in a section of formalin-fixed paraffin-embedded normal human tonsil* 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 (pH 6.0, epitope retrieval solution 1) for 20mins. The section was then incubated with ab16667, 1/200 dilution, 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. 
    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

    This image was generated from the hybridoma version.

  • Flow Cytometry - Anti-Ki67 antibody [SP6] - BSA free (ab231172)
    Flow Cytometry - Anti-Ki67 antibody [SP6] - BSA free (ab231172)

    This data was developed using the same antibody clone in a different buffer formulation (ab16667).

    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 (ab16667,1/1000) for 30 min at 22°C. The secondary antibody used was Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) secondary antibody 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.

    This image was generated from the hybridoma version.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Ki67 antibody [SP6] - BSA free (ab231172)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Ki67 antibody [SP6] - BSA free (ab231172) Image courtesy of Jing Ma by Abreview.

    This data was developed using the same antibody clone in a different buffer formulation (ab16667).

     

    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.

    This image was generated from the hybridoma version.

    See Abreview

  • Immunocytochemistry - Anti-Ki67 antibody [SP6] - BSA free (ab231172)
    Immunocytochemistry - Anti-Ki67 antibody [SP6] - BSA free (ab231172) This image is courtesy of an Abreview submitted by Peter Zentis

    This data was developed using the same antibody clone in a different buffer formulation (ab16667).

    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.

    This image was generated from the hybridoma version.

     

    See Abreview

  • Anti-Ki67 antibody [SP6] - BSA free (ab231172)
    Anti-Ki67 antibody [SP6] - BSA free (ab231172)

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
For licensing inquiries, please contact partnerships@abcam.com

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