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<strong>Figure 1 KO Validation of CDKN2A in 293 Cells</strong><br> Loading: 10 μg of 293 WT cell lysates or CDKN2A KO cell lysates. Antibodies: CDKN2A, 4211 (2 μg/mL) and beta-actin, 3779 (1 μg/mL), 1 h incubation at RT in 5% NFDM/TBST.Secondary: Goat Anti-Rabbit IgG HRP conjugate at 1:10000 dilution.
<strong>Figure 2. Immunofluorescence Validation of CDKN2A In HeLa Cells </strong><br> Immunofluorescent analysis of methanol-fixed HeLa cells labeling CDKN2A with 4211 at 20 μg/mL, followed by goat anti-rabbit IgG secondary antibody at 1/1000 dilution (red) and Hoechst staining (blue). Alpha tubulin was stained with anti-alpha tubulin antibody following by goat anti-mouse IgG secondary antibody (green). Images were captured with confocal microscopy.
<strong>Figure 3 Independent Antibody Validation (IAV) via Protein Expression Profile in Cell Lines</strong><br>Loading: 15 μg of lysates per lane.Antibodies: CDKN2A 4211 (4 μg/mL), CDKN2A 24-023 (4 μg/mL), and beta-actin (1 μg/mL), overnight incubation at 4°C in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.
<strong>Figure 4 Recombinant Protein Test</strong><br>Loading: 15 μg of lysates per lane.Antibodies: CDKN2A 4211 (2 μg/mL), overnight incubation at 4°C in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.Lane 1: Human prostate (benign hyperplasia)Lane 2: Human CDKN2A recombinant protein (arrow: monomer and dimer)
<strong>Figure 5 Western Blot Validation in Human Cell Lines</strong><br>Loading: 15 μg of lysates per lane.Antibodies: CDKN2A, 4211 (2 μg/mL), 1h incubation at RT in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.
<strong>Figure 6 Western Blot Validation in Human Normal and Cancer Tissue</strong><br>Loading: 15 μg of lysates per lane.Antibodies CDKN2A 4211 (2 μg/mL), overnight incubation at 4°C in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.Lane 1: Human breastLane 2: Human lung tumorLane 3: Human colon cancer
<strong>Figure 7 Western Blot Validation in Mouse Colon Tissue </strong><br>Loading: 15 μg of lysates per lane.Antibodies: CDKN2A 4211 (A: 1 μg/mL, B: 2 μg/mL), 1h incubation at RT in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.
<strong>Figure 8 Immunofluorescence Validation of CDKN2A in Human A431 Cells</strong><br>Immunofluorescent analysis of 4% paraformaldehyde-fixed A431 cells labeling CDKN2A with 4211 at 20 μg/mL, followed by goat anti-rabbit IgG secondary antibody at 1/500 dilution (green) and DAPI staining (blue).
<strong>Figure 9 Immunofluorescence Validation of CDKN2A in Human Colon Tissue</strong><br>Immunofluorescent analysis of 4% paraformaldehyde-fixed human colon tissue labeling CDKN2A with 4211 at 20 μg/mL, followed by goat anti-rabbit IgG secondary antibody at 1/500 dilution (green) and DAPI staining (blue).
<strong>Figure 10 Immunohistochemistry Validation of CDKN2A in Human Colon Tissue </strong><br> Immunohistochemical analysis of paraffin-embedded Human Colon Tissue using anti- CDKN2A antibody (4211) at 10 μg/ml. Tissue was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4°C. A goat anti-rabbit IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin.
<strong>Figure 11 Immunohistochemistry Validation of CDKN2A in Human Colon Tissue </strong><br> Immunohistochemical analysis of paraffin-embedded Human Colon Tissue using anti- CDKN2A antibody (4211) at 10 μg/ml. Tissue was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4°C. A goat anti-rabbit IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin.
<strong>Figure 12 Immunohistochemistry Validation of CDKN2A in Rat Colon Tissue </strong><br> Immunohistochemical analysis of paraffin-embedded Rat Colon Tissue using anti- CDKN2A antibody (4211) at 5 μg/ml. Tissue was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4°C. A goat anti-rabbit IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin.
Flow Cytometry Validation of CDKN2A in HeLa Cells 1X10^6/mL HeLa cells were intracellularly stained with 2 μg/mL of CDKN2A antibody (red) or Rabbit IgG isotype control (black) at RT for 30 min, followed by PSFlo 488 conjugated goat anti-rabbit IgG secondary antibody staining (1:1000) at RT for 30 min. Cells were fixed with 4% PFA and permeabilized with 0.1% Triton X-100 in PBS.
Figure 1 KO Validation of CDKN2A in 293 Cells
Loading: 10 μg of 293 WT cell lysates or CDKN2A KO cell lysates. Antibodies: CDKN2A, 4211 (2 μg/mL) and beta-actin, 3779 (1 μg/mL), 1 h incubation at RT in 5% NFDM/TBST.Secondary: Goat Anti-Rabbit IgG HRP conjugate at 1:10000 dilution.
IAV
KO
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CDKN2A/p16INK4a Antibody

CATALOG NUMBER: 4211

Clonality:
Polyclonal
Tested Applications:
ELISA, FC, IHC-P, WB, ICC/IF, IHC-P/IF
Host Species:
Rabbit
Species Reactivity:
Human, Mouse, Rat
Conjugate:
Unconjugated
Specifications
Host Species:
Rabbit
Species Reactivity:
Human, Mouse, Rat
Immunogen:
Anti-CDKN2A antibody (4211) was raised against a peptide corresponding to 18 amino acids near the amino terminus of human CDKN2A.
The immunogen is located within the first 50 amino acids of CDKN2A.
Conjugate:
Unconjugated
Tested Applications:
ELISA, FC, IHC-P, WB, ICC/IF, IHC-P/IF
Application Note:
WB: 1 - 4 μg/mL; IHC-P: 5 - 10 μg/mL; IF: 20 μg/mL; FC: 2 μg/mL.
Antibody validated: Western Blot in human and mouse samples; Immunohistochemistry in human and rat samples; Immunofluorescence in human samples; Flow Cytometry in human samples. All other applications and species not yet tested.
Positive Control 1:
Cat. No. 1201 - HeLa Cell Lysate
Positive Control 2:
Cat. No. 1211 - HepG2 Cell Lysate
Positive Control 3:
Cat. No. 1411 - Mouse Colon Tissue Lysate
Predicted Molecular Weight:
Predicted: 17 kDa
Observed: 17 kDa
Validation:

KO Validation (Figure 1): Anti-CDKN2A antibodies (4211) specificity was further verified by CDKN2A specific knockout. CDKN2A signal was not detected in CDKN2A knockout 293 cells as compared to that in control wild type cells.

Independent Antibody Validation in Cell lines (Figure 2) shows similar CDKN2A expression profile in human cell lines detected by two independent anti-CDKN2A antibodies that recognize different epitopes, 4211 against N-terminus domain and 8975 against the C-terminus domain.  CDKN2A proteins are detected only in 293, HeLa and HepG2 cell lines by the two independent antibodies. 

Recombinant Protein Test (Figure 3) shows that anti-CDKN2A antibody (4211) detects human recombinant CDKN2A protein that contains the amino acids of 4211 antigenic peptide.

Isoforms:
Human CDKN2A has 6 isoforms, including isoform 1 (156aa, 17kD), isoform 2 (105aa, 11kD), isoform 3 (116aa, 12kD), isoform 5 (167aa, 18kD), isoform tumor suppressor ARF (132aa, 14kD) and isoform smARF (85aa, 9kD). Mouse CDKN2A has 4 isoforms, including isoform 1 (168aa, 18kD), isoform 2 (125aa, 14kD), isoform tumor suppressor ARF (169aa, 19kD) and isoform smARF (125aa, 14kD). Rat CDKN2A has two isoforms, including isoform 1 (159aa, 17kD) and isoform ARF (160aa, 17kD). 4211 can detect only human isoforms 1, 3 and 5, mouse isoform 1 and rat isoform 1.
Purification:
CDKN2A Antibody is affinity chromatography purified via peptide column.
Clonality:
Polyclonal
Isotype:
IgG
physical-state:
Liquid
Buffer:
CDKN2A Antibody is supplied in PBS containing 0.02% sodium azide.
Concentration:
1 mg/mL
Storage Conditions:
CDKN2A antibody can be stored at 4°C, stable for up to one year. Antibodies should not be exposed to prolonged high temperatures.
Ncbi Official Symbol:
CDKN2A
Additional Names:
CDKN2A Antibody: P16, INK4, MTS1, CDK4I, CDKN2, INK4A, MTS-1, P16INK4, P16INK4A, P16-INK4A, Cyclin-dependent kinase 4 inhibitor A
Protein Accession Number:
NP_000068
Protein Gi Number:
4502749
Ncbi Gene Id Number:
1029
User Note:
Optimal dilutions for each application to be determined by the researcher.
Background:
CDKN2A Antibody: The CDKN2A locus gives rise to 2 distinct transcripts from different promoters. The transcripts have been designated p16(INK4A) and p14(ARF). This chromosomal region undergoes a number of inversions, translocations, heterozygous deletions, and homozygous deletions in a variety of malignant cell lines including those from glioma, non-small cell lung cancer, leukemia, and melanoma. Deletion of the region containing CDKN2A is found in more than half of all melanoma cell lines. Conversely, transfection of CDKN2A suppressed the growth of two independent mesothelioma cell lines, suggesting that inactivation of the CDKN2 gene is an essential step in the etiology of malignant mesotheliomas. CDKN2A induces a G1 cell cycle arrest by inhibiting the phosphorylation of the Rb protein by the cyclin-dependent kinases CDK4 and CDK6. CDKN2A is expressed as at least three distinct isoforms.
Background Reference 1:
Stone et al. Cancer Res. 1995; 55:2988-94.
Background Reference 2:
Kamb et al. Nature Genet. 1994; 8:22-6.
Background Reference 3:
Kratzke et al. J. Nat. Cancer Inst. 1995; 87:1870-5.
Background Reference 4:
Stott et al. EMBO J. 1998; 17:5001-14.

FOR RESEARCH USE ONLY

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Disclaimer:
Optimal dilutions/concentrations should be determined by the end user. The information provided is a guideline for product use. This product is for research use only.

CATALOG NUMBER:

4211

List Size:
0.02 mg, 0.1 mg

List Price:

Price range: $99.00 through $445.00

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