Description
Key facts
Detection method:Colorimetric,
Sample types:Suspension cells, Adherent cells,
Reacts with:Mouse, Human,
Assay type:Cell-based (quantitative),
Assay Platform:Microplate
Product details:
Human Cell Cycle In-Cell ELISA kit (IR) (ab139412) that uses quantitative immunocytochemistry to measure levels of Cdk2 protein phosphorylated Tyr15 and Histone H3 protein phosphorylated Ser10 levels in cultured cells. Cells are fixed in a microplate and targets of interest are detected with highly specific, well-characterized antibodies. Relative target levels are quantified using IRDye®-labeled Secondary Antibodies and IR imaging such as a LI-COR® Odyssey® or Aerius® system. Antibody signal intensity can be normalized to the total cell amount using Janus Green stain.
Plates are available in our ICE (In-Cell ELISA) Support Pack (ab111542) which can be bought seperately.
The Cdk2 (pTyr15) + Histone H3 (pSer10) In-Cell ELISA Kit (IR) (ab139412) is designed to study cell cycle effects in response to various stimuli. Monoclonal antibodies specific to Cdk2 (pTyr15) and Histone H3 (pSer10) are used in this high-throughput duplexing plate-based assay. Cdk2 (pTyr15) is elevated in G1/S phase of the cell cycle and Histone H3 (pSer10) is elevated in G2/M phase. Cyclin-dependent kinase 2 (Cdk2) is a nuclear protein kinase that functions in the G1/S phase of the cell cycle. Inhibitory phosphorylation occurs on residues Thr14 and Tyr15; activation of Cdk2 includes dephosphorylation of these residues by cdc25. Cdk2 can form a complex with Cyclin A, D or E. Phosphorylation of Cdk2 at Tyr15 indicates that a cell is at the G1/S transition.
Histone H3 is one of the four core histone proteins (H2A, H2B, H3 and H4) that pack DNA in nucleosomes. Post-translational modifications of histones include phosphorylation and acetylation and are important for chromatin assembly and gene expression. Phosphorylation of Histone H3 at Ser10 is tightly correlated with chromosome condensation during mitosis. Hence, Histone H3 pSer10 signal indicates a mitotic cell with condensed DNA.
In-Cell ELISA (ICE) technology employed to perform quantitative immunocytochemistry of cultured cells with a near-infrared fluorescent dye-labeled detector antibody. The technique generates quantitative data with specificity similar to western blotting, but with much greater quantitative precision and higher throughput due to the greater dynamic range and linearity of direct fluorescence detection and the ability to run 96 samples in parallel. Because the Cdk2 (pTyr15) antibody is a rabbit antibody and the Histone H3 (pSer10) antibody is a mouse antibody, they can be measured simultaneously in the same well using the cocktail of provided primary antibodies and the provided cocktail of IRDye®-labeled species-specific secondary antibodies when using a LI-COR infrared imager. This method rapidly fixes the cells in situ, stabilizing the in vivo levels of proteins and their post-translational modifications, and thus eliminating changes during sample handling, such as in the preparation of protein extracts. Finally, the Cdk2 (pTyr15) and Histone H3 (pSer10) signals can be normalized to cell amount, measured by the provided Janus Green whole cell stain, to further increase the assay precision.
REACH authorisation
Abcam has not and does not intend to apply for the REACH Authorisation of customers' uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.
Properties and Storage Information:
Shipped at conditions-Blue Ice, Appropriate short-term storage conditions-+4°C, Appropriate long-term storage conditions-+4°C, Storage information-+4°C
Supplementary Information:
This supplementary information is collated from multiple sources and compiled automatically.
The biological target CdK2 also known as Cyclin-Dependent Kinase 2 and Histone H3 is involved in cell cycle regulation and chromatin remodeling. CdK2 is a serine/threonine kinase with a mass of approximately 34 kDa. It is expressed in the nucleus of dividing cells. Histone H3 is one of the core histones that form the histone octamer around which DNA is wrapped and it has a mass of about 15.3 kDa. These proteins are expressed in various tissues particularly in proliferating cells throughout the body.
Biological function summary
Cyclin-Dependent Kinase 2 partners with specific cyclin proteins to drive cell cycle progression. It forms a complex with Cyclin E or Cyclin A to control the G1/S and S/G2 transitions ensuring proper DNA replication. Histone H3 undergoes various post-translational modifications like phosphorylation and acetylation which impact chromatin structure and gene expression. As part of the nucleosome it plays an important role in regulating access to genetic information.
Pathways
CdK2 and Histone H3 play key roles in the cell cycle and epigenetic regulation pathways. CdK2 interacts with Cyclin E during the transition from G1 to S phase influencing DNA replication. Cyclin A is involved in the S phase and entry into mitosis. Histone H3 modifications are central to the regulation of epigenetic pathways affecting gene expression levels and stability. These pathways integrate with numerous cellular processes and interact with several proteins including RETINOBLASTOMA (Rb) and p21 which further regulate the cell cycle.
CdK2 and Histone H3 have critical roles in cancer and neurological diseases. CdK2 overactivity can lead to uncontrolled cell proliferation contributing to various cancers. Its interaction with Cyclin E is often upregulated in tumors. Aberrant modifications of Histone H3 such as mutations in H3K27 or H3K9 link to specific cancers like gliomas. Moreover dysregulation in Histone H3 is associated with Huntington's disease involving alterations in transcriptional regulation. These connections highlight the potential for targeting CdK2 and Histone H3 in therapeutic approaches.
