Revvity, ALSU-PDNA-A500, AlphaLISA Human Phospho-DNA PKcs (Ser2056) Detection Kit, 500 Assay Points

Overview
DNA-PK, a nuclear DNA-dependent serine/threonine protein kinase, consists of two components, a DNA-dependent protein kinase, catalytic subunit (DNA PKcs) and an autoimmune antigen (KU). DNA-PKcs is inactive and relies on KU to direct it to DNA ends and trigger kinase activity where it is required for the non-homologous end joining pathway of DNA repair which rejoins double-strand breaks. DNA repair genes likely underlie many forms of cancer, and reduced expression of DNA PKcs is associated with cancers such as breast, prostate, and gastric.
The AlphaLISA SureFire Human Phospho-DNA PKcs (Ser2056) Detection Kit is a sandwich immunoassay for the quantitative detection of phospho-DNA PKcs (Ser2056) in cellular lysates, using Alpha Technology.
Formats

The HV (high volume) kit contains reagents to run 100 wells in 96-well format, using a 60 μL reaction volume.
The 500-point kit contains enough reagents to run 500 wells in 384-well format, using a 20 μL reaction volume.
The 10,000-point kit contains enough reagents to run 10,000 wells in 384-well format, using a 20 μL reaction volume.
The 50,000-point kit contains enough reagents to run 50,000 wells in 384-well format, using a 20 μL reaction volume.

AlphaLISA SureFire Ultra kits are compatible with

Cell and tissue lysates
Antibody modulators
Biotherapeutic antibodies

Alpha SureFire kits can be used for

Cellular kinase assays
Receptor activation studies
Screening
How it works
Phospho-AlphaLISA SureFire Ultra assay principle

The Phospho-AlphaLISA SureFire Ultra assay measures a protein target when phosphorylated at a specific residue.

The assay uses two antibodies which recognize the phospho epitope and a distal epitope on the targeted protein. AlphaLISA assays require two bead types: Acceptor and Donor beads. Acceptor beads are coated with a proprietary CaptSure™ agent to specifically immobilize the assay specific antibody, labeled with a CaptSure tag. Donor beads are coated with streptavidin to capture one of the detection antibodies, which is biotinylated. In the presence of phosphorylated protein, the two antibodies bring the Donor and Acceptor beads in close proximity whereby the singlet oxygen transfers energy to excite the Acceptor bead, allowing the generation of a luminescent Alpha signal. The amount of light emission is directly proportional to the quantity of phosphoprotein present in the sample.

 

Phospho-AlphaLISA SureFire Ultra two-plate assay protocol

The two-plate protocol involves culturing and treating the cells in a 96-well plate before lysis, then transferring lysates into a 384-well OptiPlate™ plate before the addition of Phospho-AlphaLISA SureFire Ultra detection reagents. This protocol permits the cells viability and confluence to be monitored. In addition, lysates from a single well can be used to measure multiple targets.

Phospho-AlphaLISA SureFire Ultra one-plate assay protocol

Detection of Phosphorylated target protein with AlphaLISA SureFire Ultra reagents can be performed in a single plate used for culturing, treatment, and lysis. No washing steps are required. This HTS designed protocol allows for miniaturization while maintaining AlphaLISA SureFire Ultra quality.
Application-Cell Signaling
Automation Compatible-Yes
Brand-AlphaLISA SureFire Ultra
Detection Modality-Alpha
Lysis Buffer Compatibility-Lysis Buffer
Molecular Modification-Phosphorylation
Product Group-Kit
Sample Volume-10 µL
Shipping Conditions-Shipped in Blue Ice
Target-DNA PKcs
Target Class-Phosphoproteins
Target Species-Human
Technology-Alpha
Therapeutic Area-Oncology
Unit Size-500 assay points