Phosphatases are a class of enzymes that play a key role in the dephosphorylation of organic compounds. Using water as a reactant, phosphatase enzymes catalyze the hydrolysis of various monophosphate esters into phosphoric acid and alcohol. These enzymes are present in both prokaryotes and eukaryotes alike, and are typically classified as either acid phosphatase or alkaline phosphatase. Acidic phosphatases function optimally in acidic pH environments, whereas alkaline phosphatases are optimally active at alkaline pH environments. Together with kinases (enzymes that catalyze the addition of phosphate groups onto molecules), phosphatase enzymes direct a form a post-translational modification that is essential for normal cell biology and pathogenesis.
Alkaline phosphatase (ALP) is an 86 kDa homodimeric protein enzyme. It contains two zinc atoms which are crucial to its catalytic function per monomer, and is optimally active at alkaline pH environments. ALP has become a highly sensitive reporter enzyme in immunoassays such as ELISA, as well as, in immunohistochemical (IHC), Northern, Southern and Western blot applications. For these applications, ALP is usually conjugated to a specific primary or secondary antibody and its activity is detected with a signal generating substrate.
AAT Bioquest provides several “mix and read” phosphatase assay kits that are compatible with HTS liquid handling instruments, as well as chromogenic and fluorogenic phosphatase substrate to quantify alkaline phosphatase activity in solutions, cell extracts, or on solid surfaces, such as PVDF membranes.