Description
Key facts
Detection method:Colorimetric,
Sample types:Plasma, Tissue Extracts, Cell culture media, Serum, Other biological fluids, Cell Lysate,
Assay type:Enzyme activity (quantitative),
Sensitivity:> 10 µU,
Range:10 - 250 µU,
Assay time:1h 10m,
Assay Platform:Microplate reader
Product details:
Alkaline Phosphatase Assay Kit (Colorimetric) ab83369 is a highly sensitive, simple, direct and HTS-ready colorimetric assay designed to measure alkaline phosphatase (ALP) activity in serum and other mammalian samples.
How the assay works
The kit is a pNPP assay; it uses p-nitrophenyl phosphate (pNPP) as a phosphatase substrate which turns yellow (λmax= 405 nm) when dephosphorylated by ALP as part of an ALP activity assay. This kit contains 10 substrate tablets providing convenience for multiple usages.
Alkaline phosphatase assay protocol summary
- Add samples and standards to wells
- Add pNPP solution to sample wells (not to standards)
- Add ALP enzyme solution to standard wells (not to samples)
- Incubate for 60 min at room temp
- Add stop solution
- Analyze with microplate reader
Related alkaline phosphatase assay products
For a fluorometric alkaline phosphatase assay kit, we recommend
ab83371
. This assay is based on the 4-Methylumbelliferyl phosphate disodium salt (MUP) substrate. Alkaline phosphatase cleaves the phosphate group of this substrate, releasing fluorescent 4-methylumbelliferyl (Ex/Em = 360nm/440nm).
For a luminescent alkaline phosphatase assay, we recommend
ab233466
. This assay is bsed on the ability of alkaline phosphatase to remove the phosphate group from D-luciferin phosphate, releasing the luciferin to be processed by luciferase.
For alkaline phosphatase staining, we recommend Alkaline Phosphatase Staining Kit
ab284936
. This kit is based on the substrate, 5-Bromo-4-chloro-3-indolyl phosphate (BCIP) which is processed by alkaline phosphatase to form a blue colored intermediate. Nitro Blue Tetrazolium (NBT) then oxidizes the intermediate to form an insoluble, dark blue dimer, which can be viewed by microscopy.
Getting the best performance from ab83369
Technical Note: Alkaline phosphatase is unlikely to be detectable in healthy urine but may be elevated in certain kidney, liver or bone disorders.
Alkaline phosphatase assay methods
pNPP assays are a class of phosphatase assays using p-nitrophenyl phosphate (pNPP) as a phosphatase substrate. They can be used to measure alkaline phosphatase, neutral phosphatase, and acid phosphatase activity. These different types of phosphatases are active respectively in alkaline, neutral, and acid assay buffers. pNPP assays include Alkaline Phosphatase Assay Kit ab83369 and Acid Phosphatase Assay Kit
ab83367
The pNPP phosphatase assay method is by far the most popular method for a colorimetric alkaline phosphatase assay for liquid samples or cell/tissue lysates.
Other notes
This product is manufactured by BioVision, an Abcam company and was previously called K412 Alkaline Phosphatase Activity Colorimetric Assay Kit. K412-500 is the same size as the 500 test size of ab83369.
The Safety Datasheet for this product has been updated for certain countries. Please check the current version in the Support and downloads section.
Properties and Storage Information:
Shipped at conditions-Blue Ice, Appropriate short-term storage conditions--20°C, Appropriate long-term storage conditions--20°C, Storage information--20°C
Supplementary Information:
This supplementary information is collated from multiple sources and compiled automatically.
Alkaline phosphatase (ALP) is an enzyme that catalyzes the hydrolysis of phosphate groups from various molecules including nucleotides and proteins. It is a homo-dimeric protein with each monomer having a mass of about 57 kDa. The enzyme is widely expressed across different tissues with high levels present in the liver bone kidney and placenta. ALP is also commonly referred to as ALP or alkaline phosphatase where it exists as various isoenzymes depending on the tissue of expression.
Biological function summary
Alkaline phosphatase is involved in essential processes such as dephosphorylation which impacts cellular communication and energy metabolism. The enzyme participates in bone mineralization by breaking down pyrophosphate an inhibitor of mineralization to facilitate calcium and phosphate deposition. It does not typically function as part of a larger complex but its activity is critical in maintaining normal physiological pH levels in various tissues. The enzyme's activity is often measured using an alkaline phosphatase assay or ALP assay valuable for assessing different condition states within the body.
Pathways
Alkaline phosphatase plays roles in bone formation and liver function pathways. It interacts with other proteins like collagen and osteocalcin in the bone mineralization pathway which is vital for healthy bone development and maintenance. Additionally in liver function ALP is linked to other enzymes like gamma-glutamyltransferase (GGT) which together help detoxify hematopoietic and other bodily functions. Alkaline phosphatase activity can influence these pathways significantly impacting health status and treatment outcomes.
Abnormal alkaline phosphatase levels often link to conditions such as Paget's disease of bone and hepatobiliary dysfunction. In Paget's disease elevated ALP indicates increased bone turnover often with increased osteocalcin activity. Meanwhile in liver disease altered ALP levels may connect with gamma-glutamyltransferase during diagnostic analyses to assess liver dysfunction or biliary obstruction. Regular monitoring of ALP activity is important for diagnosing these conditions and evaluating treatment responses.
