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[KO Validated] CDK4 Rabbit PolymAb® (A23521PM)

Tested applications:WB IHC-P IF/ICC IP ChIP ChIP-seq RIP FC FC(Intra)ELISA MeDIP Nucleotide Array DB FACS CoIP CUT&Tag meRIP InhibitionReactivity:Human, Mouse, Rat

Western blot analysis of lysates from wild type (WT) and CDK4 knockout (KO) HeLa cells using [KO Validated] CDK4 Rabbit PolymAb® (A23521PM) at 1:3000 dilution incubated overnight at 4℃.
Secondary antibody: HRP-conjugated Goat anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25 μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 30s.

Western blot analysis of various lysates using [KO Validated] CDK4 Rabbit PolymAb® (A23521PM) at 1:1000 dilution incubated overnight at 4℃.
Secondary antibody: HRP-conjugated Goat anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25 μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 30s.

ABclonal:Immunohistochemistry - [KO Validated] CDK4 Rabbit PolymAb® (A23521PM)

Immunohistochemistry analysis of paraffin-embedded Mouse testis tissue using [KO Validated] CDK4 Rabbit PolymAb® (A23521PM) at a dilution of 1:600 (40x lens). High pressure antigen retrieval performed with 0.01M Tris-EDTA Buffer (pH 9.0) prior to IHC staining.

Immunohistochemistry analysis of paraffin-embedded Human breast cancer tissue using [KO Validated] CDK4 Rabbit PolymAb® (A23521PM) at a dilution of 1:600 (40x lens). High pressure antigen retrieval performed with 0.01M Tris-EDTA Buffer (pH 9.0) prior to IHC staining.

Immunohistochemistry analysis of paraffin-embedded Rat spleen tissue using [KO Validated] CDK4 Rabbit PolymAb® (A23521PM) at a dilution of 1:600 (40x lens). High pressure antigen retrieval performed with 0.01M Tris-EDTA Buffer (pH 9.0) prior to IHC staining.

ABclonal:Immunofluorescence - [KO Validated] CDK4 Rabbit PolymAb® (A23521PM)

Confocal imaging of HeLa cells using [KO Validated] CDK4 Rabbit PolymAb® (A23521PM, dilution 1:200) followed by a further incubation with Cy3 Goat Anti-Rabbit IgG (H+L) (AS007, dilution 1:500) (Red). The cells were counterstained with α-Tubulin Mouse mAb (AC012, dilution 1:400) followed by incubation with ABflo® 488-conjugated Goat Anti-Mouse IgG (H+L) Ab (AS076, dilution 1:500) (Green). DAPI was used for nuclear staining (Blue). Objective: 100x.

Confocal imaging of paraffin-embedded Rat lung tissue using [KO Validated] CDK4 Rabbit PolymAb® (A23521PM, dilution 1:200) followed by a further incubation with Cy3 Goat Anti-Rabbit IgG (H+L) (AS007, dilution 1:500) (Red). DAPI was used for nuclear staining (Blue). High pressure antigen retrieval performed with 0.01M Citrate Buffer (pH 6.0) prior to IF staining. Objective: 40x.

ABclonal:Immunoprecipitation - [KO Validated] CDK4 Rabbit PolymAb® (A23521PM)

Immunoprecipitation of CDK4 from 300 µg extracts of HeLa cells was performed using 2 µg of [KO Validated] CDK4 Rabbit PolymAb® (A23521PM). Rabbit IgG isotype control (AC042) was used to precipitate the Control IgG sample. IP samples were eluted with 1X reducing Laemmli Buffer. The Input lane represents 10% of the total input. Western blot analysis of immunoprecipitates was conducted using [KO Validated] CDK4 Rabbit PolymAb® (A23521PM) at a dilution of 1:3000.

Overview

Product name	[KO Validated] CDK4 Rabbit PolymAb®
Catalog No.	A23521PM
Host species	Rabbit
Purification method	Affinity purification
Isotype	IgG

Background

The protein encoded by this gene is a member of the Ser/Thr protein kinase family. This protein is highly similar to the gene products of S. cerevisiae cdc28 and S. pombe cdc2. It is a catalytic subunit of the protein kinase complex that is important for cell cycle G1 phase progression. The activity of this kinase is restricted to the G1-S phase, which is controlled by the regulatory subunits D-type cyclins and CDK inhibitor p16(INK4a). This kinase was shown to be responsible for the phosphorylation of retinoblastoma gene product (Rb). Mutations in this gene as well as in its related proteins including D-type cyclins, p16(INK4a) and Rb were all found to be associated with tumorigenesis of a variety of cancers. Multiple polyadenylation sites of this gene have been reported.

Immunogen information

Immunogen	A synthetic peptide corresponding to a sequence within amino acids 204-303 of human CDK4 (NP_000066.1).
Sequence	FAEMFRRKPLFCGNSEADQLGKIFDLIGLPPEDDWPRDVSLPRGAFPPRGPRPVQSVVPEMEESGAQLLLEMLTFNPHKRISAFRALQHSYLHKDEGNPE
Gene ID
Swiss Prot
Synonyms	CMM3; PSK-J3
Calculated MW	34kDa
Observed MW	34kDa

Applications

Reactivity	Human, Mouse, Rat
Tested applications	WB IHC-P IF/ICC IP ChIP ChIP-seq RIP FC FC(Intra)ELISA MeDIP Nucleotide Array DB FACS CoIP CUT&Tag meRIP Inhibition
Recommended dilution	WB 1:1000 - 1:12000 IHC-P 1:200 - 1:800 IF/ICC 1:200 - 1:800 IP 0.5μg-4μg antibody for 200μg-400μg extracts of whole cells ELISA Recommended starting concentration is 1 μg/mL. Please optimize the concentration based on your specific assay requirements.
Storage buffer	Store at -20℃. Avoid freeze / thaw cycles. Buffer: PBS with 0.09% Sodium azide, 0.05% BSA, 50% glycerol, pH7.3.
Key application	Western blotting Immunohistochemistry Immunofluorescence Immunoprecipitation
Positive samples	Mouse lung, Rat lung, HeLa
Cellular location	Cytoplasm, Membrane, Nucleus.