Informazioni per l’ordine
Phospho-histone H3 and α-Tubulin Assay
Description:
Phospho-histone H3 and α-Tubulin Assay
Product Overview:
Millipore’s HCS216 assay provides a complete immunofluorescence-based solution for identifying and quantifying phospho-histone H3(Ser10) and α-tubulin expression in cellular imaging applications. The assay is suitable for multiple applications, including, but not limited to, the study of agents which modulate the cell cycle, analysis of microtubule-targeted drugs, cytoskeletal characterization, mitotic index determination, cancer drug screening, and in vitro toxicology studies. The highly-validated reagents provided with this kit have been specifically optimized for HCS applications, and allow the user to standardize their assays, minimize assay-to-assay variability, and to reproducibly generate images with a high signal-to-background ratio.
Background Information:
Phospho-Histone H3 (Ser10)
Histone H3 has been identified as a crucial target for cancer chemotherapy due to its important regulatory role in cell proliferation and neoplastic cell transformation. Accordingly, HCS of histone H3 phosphorylation is increasingly being used as a tool in drug discovery. HCS assays quantifying phospho-histone H3 levels have recently been utilized to characterize Aurora kinase inhibitors. Additionally, histone H3 phosphorylation has been used as a readout in cell cycle inhibitor profiling studies, illustrating the value of this approach in drug discovery applications.
α-Tubulin
Microtubules are long, filamentous, tube-shaped cytoskeletal protein polymers consisting of α- and β-tubulin heterodimers. Analysis of the dynamic assembly and disassembly of α-tubulin using cellular imaging techniques has proven invaluable in understanding the role of microtubules in cell health and disease. Microtubules are now known to be crucial in the development and maintenance of cell shape, in the transport of vesicles, mitochondria and other components throughout cells, in cell signaling, and in cell division and mitosis.
Histone H3 has been identified as a crucial target for cancer chemotherapy due to its important regulatory role in cell proliferation and neoplastic cell transformation. Accordingly, HCS of histone H3 phosphorylation is increasingly being used as a tool in drug discovery. HCS assays quantifying phospho-histone H3 levels have recently been utilized to characterize Aurora kinase inhibitors. Additionally, histone H3 phosphorylation has been used as a readout in cell cycle inhibitor profiling studies, illustrating the value of this approach in drug discovery applications.
α-Tubulin
Microtubules are long, filamentous, tube-shaped cytoskeletal protein polymers consisting of α- and β-tubulin heterodimers. Analysis of the dynamic assembly and disassembly of α-tubulin using cellular imaging techniques has proven invaluable in understanding the role of microtubules in cell health and disease. Microtubules are now known to be crucial in the development and maintenance of cell shape, in the transport of vesicles, mitochondria and other components throughout cells, in cell signaling, and in cell division and mitosis.
Key Applications:
- in vitro Toxicology Assays
- Immunocytochemistry
- High Content Analysis
Application Notes:
Cell Cycle
Usage Statement:
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
Kit or Assay Type:
High-Content Screening
Configuration:
Sufficient reagents for 5 X 96 well plates
Components:
- 1. Rabbit Anti-Phospho-Histone H3(Ser10) HCS Primary Antibody, 100X
- 2. HCS Secondary Antibody (donkey anti-rabbit IgG, FITC conjugate), 200X
- 3. Mouse Anti-α-Tubulin HCS Primary Antibody, 100X
- 4. HCS Secondary Antibody (donkey anti-mouse IgG, Cy3 conjugate), 200X
- 5. Hoechst HCS Nuclear Stain, 200X
- 6. HCS Fixation Solution with Phenol Red, 2X
- 7. HCS Immunofluorescence Buffer, 1X
- 8. HCS Wash Buffer, 1X
- 9. Etoposide (S/G2 Arrest Control Compound), 250X
- 10. Paclitaxel (G2/M Arrest Control Compound), 250X
- 11. Nocodazole (G2/M Arrest Control Compound), 250X
- 12. Vinblastine Sulfate (G2/M Arrest Control Compound), 250X
- 13. DMSO for Compound Serial Dilution
- 14. Compound Dilution Buffer
- 15. Plate Sealers
Storage Conditions:
Store kit components under the conditions indicated on the labels. Discard any remaining reagents after 6 months.
Detection Methods:
Fluorescent
Materials Required but Not Delivered:
1. Sterile, tissue culture-treated black/clear bottom microplates suitable for High-Content Imaging.
2. Cell-type for assay, e.g., HeLa (human cervical adenocarcinoma, ATCC #CCL-2) or A549 (human lung carcinoma, ATCC #CCL-185).
3. Tissue culture instruments/supplies (including 37ºC incubator, growth media, flasks/plates, detachment buffer, etc.) for cell type of interest.
4. HCS imaging/analysis system, e.g., GE Healthcare IN Cell Analyzer 1000 with Investigator software. System must be equipped with beam-splitters and filters capable of reading emission spectra in the blue, green and red ranges.
2. Cell-type for assay, e.g., HeLa (human cervical adenocarcinoma, ATCC #CCL-2) or A549 (human lung carcinoma, ATCC #CCL-185).
3. Tissue culture instruments/supplies (including 37ºC incubator, growth media, flasks/plates, detachment buffer, etc.) for cell type of interest.
4. HCS imaging/analysis system, e.g., GE Healthcare IN Cell Analyzer 1000 with Investigator software. System must be equipped with beam-splitters and filters capable of reading emission spectra in the blue, green and red ranges.