CellSimple™ Cleaved Caspase-3 and Phospho-Histone H3 Antibody Assay Kit is a fluorescent assay designed for use with the CellSimple™ Cell Analyzer. The kit includes Cleaved Caspase-3 (Asp175) (D3E9) Rabbit mAb (PE Conjugate) #12768 and Phospho-Histone H3 (Ser10) (D2C8) XP® Rabbit mAb (Alexa Fluor® 488) #3465. The kit allows for the quick and easy assessment of the levels of mitotic cells undergoing apoptosis.
The conjugated antibodies in this kit are supplied in PBS (pH 7.2), less than 0.1% sodium azide and 2 mg/ml BSA. Store at 4°C. Do not aliquot the antibodies. Protect from light. Do not freeze. The remaining components of the kit should be stored at the recommended temperature upon receipt. All components in this kit are stable for at least 12 months when stored at the recommended temperature and left unused.
Antibodies provided in the CellSimple™ Cleaved Caspase-3 and Phospho-Histone H3 Antibody Assay Kit detect endogenous levels of their respective target.
The CellSimple™ Cell Analyzer is a benchtop instrument that utilizes a disposable thin-film cassette and a combination of a 488 nm laser, two photomultiplier tubes (525/45 nm and 561 nm LP filters), Coulter Principle-based cell measurements, and on-board software to provide easy-to-run applications and data analysis. Data acquisition occurs within approximately 10 seconds per test. The instrument relies on disposable cassettes for sample handling, which alleviates the need for flow cell cleaning and fluidics maintenance and the instrument is small enough to be portable between the lab bench and the hood. Applications include quantitative assessments of cell viability, apoptosis, other labeled antibody markers and single and multiplexed bead-based assays for protein and cellular analysis.
Caspase-3 (CPP-32, Apoptain, Yama, SCA-1) is a critical executioner of apoptosis, as it is either partially or totally responsible for the proteolytic cleavage of many key proteins, such as the nuclear enzyme poly (ADP-ribose) polymerase (PARP) (1). Activation of caspase-3 requires proteolytic processing of its inactive zymogen into activated p17 and p12 fragments. Cleavage of caspase-3 requires the aspartic acid residue at the P1 position (2). Modulation of chromatin structure plays an important role in the regulation of transcription in eukaryotes. The nucleosome, made up of DNA wound around eight core histone proteins (two each of H2A, H2B, H3, and H4), is the primary building block of chromatin (3). The amino-terminal tails of core histones undergo various post-translational modifications, including acetylation, phosphorylation, methylation, and ubiquitination (4-6). These modifications occur in response to various stimuli and have a direct effect on the accessibility of chromatin to transcription factors and, therefore, gene expression (7). Phosphorylation at Ser10 of histone H3 is tightly correlated with chromosome condensation during both mitosis and meiosis (8).
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc. CellSimple is a trademark of Cell Signaling Technology, Inc. XP is a registered trademark of Cell Signaling Technology, Inc. Alexa Fluor is a registered trademark of Life Technologies Corporation. This product is provided under an intellectual property license from Life Technologies Corporation. The transfer of this product is contingent on the buyer using the purchased product solely in research, including use with HCS or other automated imaging applications but excluding use in combination with DNA microarrays. The buyer must not sell or otherwise transfer this product or its components for (a) diagnostic, therapeutic or prophylactic purposes; (b) testing, analysis or screening services, or information in return for compensation on a per-test basis; (c) manufacturing or quality assurance or quality control, or (d) resale, whether or not resold for use in research. For information on purchasing a license to this product for purposes other than as described above, contact Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, CA 92008 USA or firstname.lastname@example.org.