Phosphorylation Assay Kits

Protein phosphorylation and dephosphorylation is a closely controlled biochemical process that responds to a range of internal and external stimuli. Phosphorylation status influences a protein's subcellular distribution, interactions with other proteins, tertiary and quaternary structure, and overall activity. Since phosphorylation is a dynamic process, quantifying these phosphorylation events is crucial. Phosphorylation assay kits from Creative Biolabs allow for the rapid, precise, and quantitative detection of cellular proteins, especially those phosphoproteins from cells that use no-wash fluorescence technology. It can be used for both fundamental research and drug development in a number of areas, such as metabolism, cancer, immunology, and the central nervous system. It can also be used for a wide range of additional purposes, such as the measurement of GPCRs, kinases, and cytokines, among other biomarkers.

Features of Phosphorylation Assay Kits

• Easy to use: no-wash fluorescence technology.
• High sensitivity: able to identify target at femtogram levels.
• Savings in terms of time: effort, quick and precise outcomes.
• Compatible with a wide range of sample types: serum, plasma, CSF, cell lysates, tissue lysates, and cell culture supernatants.
• Wide range of applications: fundamental research, lead optimization, HTS screening, target discovery, and validation.

Applications of Phosphorylation Assay Kits

• Quantitative capabilities

It would be very helpful to be able to determine with certainty if a certain protein is phosphorylated using a simple and uncomplicated technique. Our phosphorylation test kits were homogenous and hyperbranched, with exceptional yield, quick binding kinetics, and outstanding selectivity for phosphoproteins.

• In vitro kinase assays and inhibitor screening

General phosphorylation can be found using the phosphorylation assay kit. It can be used to look at kinase or phosphatase activity in addition to detecting and measuring a protein's degree of phosphorylation. Analyzing the phosphorylation of serine/threonine substrates would be very helpful with phosphorylation assays. Thirty tyrosine kinases have emerged as important therapeutic targets, and each year a large number of novel inhibitors are evaluated. These screenings entail the creation of phosphosite-specific antibodies that accommodate each known site of phosphorylation and the immobilization of ELISA plates with known substrates.

Fig.1 Measurement of Syk activity inhibition.¹

• Detection of endogenous phosphorylation

The identification and quantification of alterations occurring from in vivo phosphorylation continue to provide significant challenges. Through the establishment of a stable G1 state, until cells receive the proper signal to divide, Cdh1 plays a crucial function in the regulation of the cell cycle. Cdh1 can direct particular substrates to the anaphase-promoting complex (APC) for ubiquitination and degradation when it is in an active, dephosphorylated form in G1. After cells commit to dividing in the late G1 phase, Cdk phosphorylates Cdh1, preventing it from attracting substrates to the APC and permitting the build-up of proteins that support the S and M phases that follow. The phosphatase Cdc14 dephosphorylates Cdh1 in late M, encouraging mitotic exit and starting a new cycle. The novel phosphorylation events on endogenous proteins can be assessed by gathering asynchronous developing cells, which are primarily S, G2, and M phase cells with phosphorylated Cdh1, and cells arrested in the G1 phase.

Fig.2 Detection of Cdh1 phosphorylation at different stages.¹

Reference

1. Iliuk A.; et al. Phosphorylation assay based on multifunctionalized soluble nanopolymer. Anal Chem. 2011, 83(7): 2767-74.