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Knockout Tested Rabbit Recombinant Monoclonal p27 KIP 1 antibody. Carrier free. Suitable for IP, WB, ICC/IF, Flow Cyt (Intra), IHC-P and reacts with Rat, Human, Mouse samples. Cited in 11 publications.


Images

Immunoprecipitation - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (AB206927), expandable thumbnail
  • Western blot - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (AB206927), expandable thumbnail
  • Western blot - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (AB206927), expandable thumbnail
  • Flow Cytometry (Intracellular) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (AB206927), expandable thumbnail
  • Flow Cytometry (Intracellular) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (AB206927), expandable thumbnail

Publications

Key facts

Isotype

IgG

Host species

Rabbit

Storage buffer

pH: 7.2 - 7.4
Constituents: PBS

Form

Liquid

Clonality

Monoclonal

Immunogen

  • The exact immunogen used to generate this antibody is proprietary information.

Reactivity data

Select an application
Product promiseTestedExpectedPredictedNot recommended
IPWBICC/IFFlow Cyt (Intra)IHC-P
Human
Expected
Tested
Expected
Expected
Tested
Mouse
Expected
Expected
Tested
Tested
Expected
Rat
Tested
Expected
Expected
Expected
Expected

Tested
Tested

Species

Rat

Dilution info

-

Notes

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

Expected
Expected

Species

Human, Mouse

Dilution info

Use at an assay dependent concentration.

Notes

-

Tested
Tested

Species

Human

Dilution info

-

Notes

-

Expected
Expected

Species

Rat, Mouse

Dilution info

Use at an assay dependent concentration.

Notes

-

Tested
Tested

Species

Mouse

Dilution info

-

Notes

-

Expected
Expected

Species

Rat, Human

Dilution info

Use at an assay dependent concentration.

Notes

-

Tested
Tested

Species

Mouse

Dilution info

-

Notes

ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

Expected
Expected

Species

Rat, Human

Dilution info

Use at an assay dependent concentration.

Notes

-

Tested
Tested

Species

Human

Dilution info

-

Notes

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

Expected
Expected

Species

Rat, Mouse

Dilution info

Use at an assay dependent concentration.

Notes

-

Associated Products

Select an associated product type

15 products for Alternative Product

3 products for Alternative Version

Target data

Function

Important regulator of cell cycle progression. Inhibits the kinase activity of CDK2 bound to cyclin A, but has little inhibitory activity on CDK2 bound to SPDYA (PubMed:28666995). Involved in G1 arrest. Potent inhibitor of cyclin E- and cyclin A-CDK2 complexes. Forms a complex with cyclin type D-CDK4 complexes and is involved in the assembly, stability, and modulation of CCND1-CDK4 complex activation. Acts either as an inhibitor or an activator of cyclin type D-CDK4 complexes depending on its phosphorylation state and/or stoichometry.

Alternative names

Cyclin-dependent kinase inhibitor 1B, Cyclin-dependent kinase inhibitor p27, p27Kip1, CDKN1B, p27, KIP1

Recommended products

Knockout Tested Rabbit Recombinant Monoclonal p27 KIP 1 antibody. Carrier free. Suitable for IP, WB, ICC/IF, Flow Cyt (Intra), IHC-P and reacts with Rat, Human, Mouse samples. Cited in 11 publications.

Alternative names

Cyclin-dependent kinase inhibitor 1B, Cyclin-dependent kinase inhibitor p27, p27Kip1, CDKN1B, p27, KIP1

Key facts

Isotype

IgG

Form

Liquid

Clonality

Monoclonal

Immunogen
  • The exact immunogen used to generate this antibody is proprietary information.
Carrier free

Yes

Clone number

Y236

Purification technique

Affinity purification Protein A

Specificity

This antibody recognises p27(Kip1). The rat recommendation is based on the WB results. We do not guarantee IHC-P for rat.

Dissociation constant

2.1 x 10-11 M

Concentration
Loading...

Storage

Shipped at conditions

Blue Ice

Appropriate long-term storage conditions

+4°C

Storage information

Do Not Freeze

Notes

ab206927 is the carrier-free version of Anti-p27 KIP 1 antibody [Y236] ab32034.

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

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 batch production

For more information, read more on recombinant antibodies.

Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.

This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with 1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.

Activity summary

The protein known as p27 KIP1 also referred to as p27 CDKN1B or KIP1 is a member of the kinase inhibitory protein family. It plays an important role in cell cycle regulation by inhibiting cyclin-dependent kinases (CDKs). p27 KIP1 weighs approximately 27 kDa and can be found in various tissues where it regulates cell proliferation. The protein functions as a suppressor by binding to cyclin-CDK complexes preventing the transition from G1 phase to S phase in the cell division cycle.

Biological function summary

The function of p27 KIP1 involves its role in controlling cell growth and division. It achieves this by becoming a part of larger protein complexes involving CDKs and cyclins. By directly interacting with these complexes p27 KIP1 modulates cell cycle progression therefore acting as a brake on cellular proliferation. The protein is important in maintaining proper cell cycle checkpoints and preventing uncontrolled cell growth which is essential for normal cellular functioning.

Pathways

The involvement of p27 KIP1 centers on cell cycle regulation and signaling pathways such as the PI3K/AKT pathway. Its interaction with CDKs and cyclins situates it within the core mechanisms that determine cell division timing. p27 KIP1 operates alongside other proteins like cyclin D and CDK4/6 fitting into the regulatory intricacies of these pathways. Proper functioning of these pathways ensures cellular homeostasis and prevents the development of oncogenic processes.

Associated diseases and disorders

Dysfunction of p27 KIP1 has links to cancer and neurodegenerative diseases. Low levels or mutations can lead to uncontrolled cell proliferation contributing to the development and progression of cancers such as breast cancer. p27 KIP1's role in neurodegenerative diseases involves its regulation of neuronal cell cycle re-entry with abnormalities potentially exacerbating conditions like Alzheimer's disease. In these contexts its interaction with proteins such as cyclin E and CDK2 becomes particularly relevant in understanding disease mechanisms.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
Terms & Conditions.

16 product images

  • Immunoprecipitation - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Immunoprecipitation - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Anti-p27 KIP 1 antibody [Y236] ab32034 (purified) at 1:20 dilution (2μg) immunoprecipitating p27 KIP 1 in MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate.

    Lane 1 (input): MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate 10μg
    Lane 2 (+): Anti-p27 KIP 1 antibody [Y236] ab32034 & MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate
    Lane 3 (-): Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of Anti-p27 KIP 1 antibody [Y236] ab32034 in MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate

    For western blotting, VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) was used for detection at 1:1000 dilution.
    Blocking and diluting buffer: 5% NFDM/TBST.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

    All lanes: Immunoprecipitation - Anti-p27 KIP 1 antibody [Y236] (Anti-p27 KIP 1 antibody [Y236] ab32034)

    Predicted band size: 22 kDa

  • Western blot - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Western blot - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    False colour image of Western blot: Anti-p27 KIP 1 antibody [Y236] staining at 1/5000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-p27 KIP 1 antibody [Y236] ab32034 was shown to bind specifically to p27 KIP 1. A band was observed at 28 kDa in wild-type RAW 264.7 cell lysates with no signal observed at this size in CDKN1B knockout cell line Mouse CDKN1B (p27 KIP 1) knockout RAW 264.7 cell line ab281619 (knockout cell lysate Mouse CDKN1B (p27 KIP 1) knockout RAW 264.7 cell lysate ab282970). To generate this image, wild-type and CDKN1B knockout RAW 264.7 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/20000 dilution.

    All lanes: Western blot - Anti-p27 KIP 1 antibody [Y236] (Anti-p27 KIP 1 antibody [Y236] ab32034) at 1/5000 dilution

    Lane 1: Wild-type RAW 264.7 cell lysate at 20 µg

    Lane 2: CDKN1B knockout RAW 264.7 cell lysate at 20 µg

    Performed under reducing conditions.

    Predicted band size: 22 kDa

    Observed band size: 28 kDa

  • Western blot - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Western blot - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    This WB data was generated using the same anti-p27 KIP 1 antibody clone, Y236, in a different buffer formulation (cat# Anti-p27 KIP 1 antibody [Y236] ab32034).

    Lane 1: Wild type HAP1 whole cell lysate (20 μg)
    Lane 2: CDKN1B knockout HAP1 whole cell lysate (20 μg)
    Lane 3: HeLa whole cell lysate (20 μg)
    Lane 4: MCF7 whole cell lysate (20 μg)

    Lanes 1 - 4: Merged signal (red and green). Green - Anti-p27 KIP 1 antibody [Y236] ab32034 observed at 30 kDa. Red - loading control, Anti-GAPDH antibody [6C5] - Loading Control ab8245, observed at 37 kDa.

    Unpurified Anti-p27 KIP 1 antibody [Y236] ab32034 was shown to specifically react with CDKN1B in wild-type HAP1 cells. No band was observed when CDKN1B knockout samples were used. Wild-type and CDKN1B knockout samples were subjected to SDS-PAGE. Anti-p27 KIP 1 antibody [Y236] ab32034 and Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/1000 and 1/10000 respectively. Blots were developed with 800CW Goat anti Rabbit and 680CW Goat anti Mouse secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

    All lanes: Western blot - Anti-p27 KIP 1 antibody [Y236] (Anti-p27 KIP 1 antibody [Y236] ab32034)

    Predicted band size: 22 kDa

  • Flow Cytometry (Intracellular) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Flow Cytometry (Intracellular) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Overlay histogram showing HAP1 wildtype (green line) and HAP1-CDKN1B knockout cells (red line) stained with Anti-p27 KIP 1 antibody [Y236] ab32034. 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 (Anti-p27 KIP 1 antibody [Y236] ab32034, 1μg/ml) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) presorbed (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/2000 dilution for 30 min at 22°C. A rabbit IgG isotype control antibody (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) was used at the same concentration and conditions as the primary antibody (HAP1 wildtype - black line, HAP1-CDKN1B 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 antibody can also be used in HAP1 cells fixed with 4%PFA (10 min), permeabilized with 0.1% PBS-Triton X-100 for 15 min under the same conditions.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • Flow Cytometry (Intracellular) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Flow Cytometry (Intracellular) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Intracellular Flow Cytometry analysis of MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling p27 KIP 1 with purified Anti-p27 KIP 1 antibody [Y236] ab32034 at 1/50 dilution (10 μg/ml) (red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488) secondary antibody was used at 1/2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • Immunocytochemistry/ Immunofluorescence - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Immunocytochemistry/ Immunofluorescence - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Immunocytochemistry/Immunofluorescence analysis of HeLa (human cervix adenocarcinoma) cells labelling p27 KIP 1 (green) with purified Anti-p27 KIP 1 antibody [Y236] ab32034 at 1/500. Cells were fixed with 4% Paraformaldehyde and permeabilized with 0.1% Triton X-100. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077, Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. Nuclei were counterstained with DAPI (blue).

    Secondary Only Control: PBS was used instead of the primary antibody as the negative control.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Immunohistochemical analysis of paraffin-embedded human breast carcinomaThis data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • Immunocytochemistry/ Immunofluorescence - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Immunocytochemistry/ Immunofluorescence - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Unpurified Anti-p27 KIP 1 antibody [Y236] ab32034 staining p27 KIP1 in MCF7 cells treated with NS 398 (NS 398, COX-2 inhibitor ab120295), by ICC/IF. Increase in p27 KIP1 expression correlates with increased concentration of NS 398, as described in literature.
    The cells were incubated at 37°C for 6h in media containing different concentrations of NS 398, COX-2 inhibitor ab120295 (NS 398) in DMSO, fixed with 4% formaldehyde for 10 minutes at room temperature and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with Anti-p27 KIP 1 antibody [Y236] ab32034 (1/100 dilution) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 goat anti-rabbit polyclonal antibody (Goat Anti-Rabbit IgG H&L (DyLight® 488) preadsorbed ab96899) at 1/250 dilution was used as the secondary antibody.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • Flow Cytometry (Intracellular) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Flow Cytometry (Intracellular) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Unpurified Anti-p27 KIP 1 antibody [Y236] ab32034 staining p27 KIP 1 in the human cell line MCF-7 (human breast carcinoma) by intracellular flow cytometry. Cells were fixed with 4% paraformaldehyde, permeabilized with 90% methanol and the sample was incubated with the primary antibody at a dilution of 1/20. A goat anti rabbit IgG (Alexa Fluor® 488) at a dilution of 1/2000 was used as the secondary antibody.

    Isoytype control: Rabbit monoclonal IgG (Black)

    Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue)

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Unpurified Anti-p27 KIP 1 antibody [Y236] ab32034 showing positive staining in Colonic adenocarcinoma tissue.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Unpurified Anti-p27 KIP 1 antibody [Y236] ab32034 showing positive staining in Glioma tissue.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Unpurified Anti-p27 KIP 1 antibody [Y236] ab32034 showing positive staining in Ovarian carcinoma tissue.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Unpurified Anti-p27 KIP 1 antibody [Y236] ab32034 showing positive staining in Stomach adenocarcinoma tissue.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-p27 KIP 1 antibody [Y236] ab32034).

  • OI-RD Scanning - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    OI-RD Scanning - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody.
    Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    This IHC data was generated using the same anti-p27 KIP 1 antibody clone, Y236, in a different buffer formulation (cat# Anti-p27 KIP 1 antibody [Y236] ab32034).

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human cervical carcinoma tissue sections labeling p27 KIP 1 with Purified Anti-p27 KIP 1 antibody [Y236] ab32034 at 1:50 dilution (10.4 μg/ml). Heat mediated antigen retrieval was performed using Antigen Retrieval Buffer (100X Tris-EDTA Buffer, pH 9.0) ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.

  • Western blot - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927), expandable thumbnail

    Western blot - Anti-p27 KIP 1 antibody [Y236] - BSA and Azide free (ab206927)

    Western blot: Anti-CDKN1B antibody [Y236] (Anti-p27 KIP 1 antibody [Y236] ab32034) staining at 1/5000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, Anti-p27 KIP 1 antibody [Y236] ab32034 was shown to bind specifically to CDKN1B. A band was observed at 30 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in CDKN1B knockout cell line. To generate this image, wild-type and CDKN1B knockout HCT 116 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

    All lanes: Western blot - Anti-p27 KIP 1 antibody [Y236] (Anti-p27 KIP 1 antibody [Y236] ab32034) at 1/5000 dilution

    Lane 1: Wild-type HCT 116 cell lysate at 20 µg

    Lane 2: CDKN1B knockout HCT 116 cell lysate at 20 µg

    Lane 3: Wild-type HAP1 cell lysate at 20 µg

    Lane 4: CDKN1B knockout HAP1 cell lysate at 20 µg

    Secondary

    All lanes: Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

    Performed under reducing conditions.

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