Control Lysate
(HEK293 cells were grown in a T175 flask at 37 degree C, 5%CO2, in DMEM with 10% FBS and cultured to 90% confluence. The medium was removed and cells were stimulated with1 uM of PMA for 30 min at 37 degree C. The medium was removed and cells were lysed with 4 mL of 1X Lysis Buffer #1 containing 1 mM NaF and 2 mM Na3VO4, on an orbital shaker (400 rpm) for 30 min at room temperature. Soluble supernatants were collected after a 10 min centrifugation and tested using the two-plate assay protocol.)
BAD assay kit
Phospho-BAD (S112) TR-FRET Cellular Assay Kit
Predicted: Mouse, Rat, Monkey
Predicted: Mouse, Rat, Monkey
Acc-phospho-BAD (20 uL)
BAD control lysate (100 uL)
5X Lysis Buffer #1 (1 mL)
10X TR-FRET Detection buffer (50 uL)
Control Lysate
(HEK293 cells were grown in a T175 flask at 37 degree C, 5%CO2, in DMEM with 10% FBS and cultured to 90% confluence. The medium was removed and cells were stimulated with1 uM of PMA for 30 min at 37 degree C. The medium was removed and cells were lysed with 4 mL of 1X Lysis Buffer #1 containing 1 mM NaF and 2 mM Na3VO4, on an orbital shaker (400 rpm) for 30 min at room temperature. Soluble supernatants were collected after a 10 min centrifugation and tested using the two-plate assay protocol.)
Control Lysate
(HEK293 cells were grown in a T175 flask at 37 degree C, 5%CO2, in DMEM with 10% FBS and cultured to 90% confluence. The medium was removed and cells were stimulated with1 uM of PMA for 30 min at 37 degree C. The medium was removed and cells were lysed with 4 mL of 1X Lysis Buffer #1 containing 1 mM NaF and 2 mM Na3VO4, on an orbital shaker (400 rpm) for 30 min at room temperature. Soluble supernatants were collected after a 10 min centrifugation and tested using the two-plate assay protocol.)
Testing Data
(MCF-7 cells were seeded at 50,000 cells/well and cultured overnight at 37 degree C, 5% CO2, in DMEM with 10% FBS. Cells were then treated at 37oC with either PMA for 30 min (stimulation) or with UCN 01 for 30 min then 5 uM PMA (inhibition). The media was removed from the wells, and cells were lysed with 50 uL/well of 1X Lysis Buffer #1 containing 1 mM NaF and 2 mM Na3VO4, on an orbital shaker (400 rpm) for 30 min at room temperature. The lysates were then analyzed for phospho-BAD (S112) using the two-plate assay protocol. The TR-FRET signal was recorded at 665 and 615 nm (EnVision lamp excitation) after a 4 h incubation period at room temperature.)
Testing Data
(MCF-7 cells were seeded at 50,000 cells/well and cultured overnight at 37 degree C, 5% CO2, in DMEM with 10% FBS. Cells were then treated at 37oC with either PMA for 30 min (stimulation) or with UCN 01 for 30 min then 5 uM PMA (inhibition). The media was removed from the wells, and cells were lysed with 50 uL/well of 1X Lysis Buffer #1 containing 1 mM NaF and 2 mM Na3VO4, on an orbital shaker (400 rpm) for 30 min at room temperature. The lysates were then analyzed for phospho-BAD (S112) using the two-plate assay protocol. The TR-FRET signal was recorded at 665 and 615 nm (EnVision lamp excitation) after a 4 h incubation period at room temperature.)
A superior alternative to ELISA
Features :: Benefits
• TR-FRET technology :: Proven, sensitive and robust assay technology
• Homogeneous, add-and-read assay :: No wash, no separation steps
• < 1 hour bench time :: Faster, easier than ELISA
• Adherent or suspension cells :: Higher flexibility
• Validated assays :: Results you can trust
• Small sample volume (15uL) :: Save precious samples
• Compatible with low-volume 96-well and 384-well plates : Higher throughput
• Signal stable over time :: Flexibility to read plate
• Kits available in 100 and 500 assay points :: Cost-effective and flexible
• Kits available for phospho, total and phospho+total protein :: Allows multiple protein measurements in one plate from one sample
NCBI and Uniprot Product Information
NCBI Description
The protein encoded by this gene is a member of the BCL-2 family. BCL-2 family members are known to be regulators of programmed cell death. This protein positively regulates cell apoptosis by forming heterodimers with BCL-xL and BCL-2, and reversing their death repressor activity. Proapoptotic activity of this protein is regulated through its phosphorylation. Protein kinases AKT and MAP kinase, as well as protein phosphatase calcineurin were found to be involved in the regulation of this protein. Alternative splicing of this gene results in two transcript variants which encode the same isoform. [provided by RefSeq, Jul 2008]
Uniprot Description
BAD: a proapoptotic member of the Bcl-2 family. Displaces Bax from binding to Bcl-2 and Bcl-xL, resulting in cell death. Survival factors such as IL-3 can inhibit the apoptotic activity of Bad inducing the phosphorylation of Bad by Akt and p90RSK. 14-3-3 proteins bind phosphorylated Bad, inhibiting its binding to Bcl-2 and Bcl-xL. Phosphorylation by mitochondria-anchored PKA in the BH3 domain can block the dimerization of Bad and Bcl-xL.
Protein type: Apoptosis
Chromosomal Location of Human Ortholog: 11q13.1
Cellular Component: mitochondrial outer membrane; mitochondrion; cytosol
Molecular Function: protein kinase B binding; protein binding; protein heterodimerization activity; phospholipid binding; caspase activator activity; protein kinase binding; lipid binding; protein phosphatase 2B binding
Biological Process: response to oleate; nerve growth factor receptor signaling pathway; positive regulation of apoptosis; positive regulation of proteolysis; apoptosis; response to glucocorticoid stimulus; positive regulation of caspase activity; cellular process regulating host cell cycle in response to virus; glucose homeostasis; response to estradiol stimulus; positive regulation of apoptosis by virus; response to glucose stimulus; pore complex biogenesis; positive regulation of autophagy; positive regulation of glucokinase activity; response to drug; caspase activation; epidermal growth factor receptor signaling pathway; release of cytochrome c from mitochondria; fibroblast growth factor receptor signaling pathway; phosphoinositide-mediated signaling; cytokine and chemokine mediated signaling pathway; suppression by virus of host apoptosis; ADP metabolic process; positive regulation of insulin secretion; response to testosterone stimulus; response to amino acid stimulus; ATP metabolic process; glucose catabolic process; response to ethanol; induction of apoptosis via death domain receptors; DNA damage response, signal transduction resulting in induction of apoptosis; response to hydrogen peroxide; positive regulation of T cell differentiation; positive regulation of B cell differentiation; innate immune response; regulation of mitochondrial membrane permeability; response to calcium ion; response to progesterone stimulus; positive regulation of epithelial cell proliferation
Research Articles on BAD
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Product Notes
The Human Predicted: Mouse, Rat, Monkey BAD bad (Catalog #AAA298009) is an Assay Kit and is intended for research purposes only. The product is available for immediate purchase. The AAA298009 Assay Kit recognizes Human Predicted: Mouse, Rat, Monkey BAD. AAA Biotech's BAD can be used in a range of immunoassay formats including, but not limited to, Cell-Based Assays. Researchers should empirically determine the suitability of the BAD bad for an application not listed in the data sheet. Researchers commonly develop new applications and it is an integral, important part of the investigative research process. It is sometimes possible for the material contained within the vial of "BAD, Assay Kit" to become dispersed throughout the inside of the vial, particularly around the seal of said vial, during shipment and storage. We always suggest centrifuging these vials to consolidate all of the liquid away from the lid and to the bottom of the vial prior to opening. Please be advised that certain products may require dry ice for shipping and that, if this is the case, an additional dry ice fee may also be required.Precautions
All products in the AAA Biotech catalog are strictly for research-use only, and are absolutely not suitable for use in any sort of medical, therapeutic, prophylactic, in-vivo, or diagnostic capacity. By purchasing a product from AAA Biotech, you are explicitly certifying that said products will be properly tested and used in line with industry standard. AAA Biotech and its authorized distribution partners reserve the right to refuse to fulfill any order if we have any indication that a purchaser may be intending to use a product outside of our accepted criteria.Disclaimer
Though we do strive to guarantee the information represented in this datasheet, AAA Biotech cannot be held responsible for any oversights or imprecisions. AAA Biotech reserves the right to adjust any aspect of this datasheet at any time and without notice. It is the responsibility of the customer to inform AAA Biotech of any product performance issues observed or experienced within 30 days of receipt of said product. To see additional details on this or any of our other policies, please see our Terms & Conditions page.Item has been added to Shopping Cart
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