IMAP Evaluation Demo Kit
Intended Use
The IMAP Evaluation Demo Kit allows one to set up calibration curves that can be used in conjunction with kinase, phosphatase and phosphodiesterase assays to determine percent phosphorylation in experimental samples. The calibration curves can also demonstrate microplate reader performance for FP and TR-FRET modes.
Principle of the Assay
IMAP technology is based on the specific, covalent-coordinate, high-affinity interaction of trivalent metal containing nanoparticles with phosphogroups. These phosphogroups can be free, or linked to serines, threonines, tyrosines, or other molecules, making IMAP a generic platform to assess kinase, phosphatase and phosphodiesterase activity. This basic principle has been used in the IMAP Binding System using both fluorescence polarization (FP) and time-resolved fluorescence resonance energy transfer (TR-FRET) as a read-out. In a microwell assay format, fluorescently labeled peptides are phosphorylated in a kinase reaction. Addition of the IMAP Binding System stops the kinase reaction and specifically binds the phosphorylated substrates. Phosphorylation and subsequent binding of the substrate to the beads can be detected either by FP (Figure 1) or TR-FRET (Figure 2).
Kit Contents
- IMAP Progressive Binding Buffer A
- IMAP Progressive Binding Buffer B
- IMAP Reaction Buffer containing BSA
- IMAP Reaction Buffer containing Tween-20
- IMAP Progressive Binding Reagent
- Tb-Donor
- 5FAM-PKAtide
- 5FAM-PhosphoPKAtide
When used as suggested, each kit provides sufficient reagents for 800 data points in 384-well plate format.
Assay Performance
Ordering Information
Go to the IMAP Ordering Information page.
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