What is the purpose of the BCIP reagent?
What is the purpose of the BCIP reagent?
The Alkaline Phosphatase Color Development Reagents, BCIP (5-bromo-4-chloro-3-indoyl phosphate p-toluidine salt) and NBT (p-nitro blue tetrazolium chloride), are used for detecting antigens bound to nitrocellulose or other membranes.
How do you make a BCIP?
BCIP is sold as a disodium salt or as a p-toluidine salt. To prepare 50 mg/mL BCIP disodium salt, dissolve 0.5 g in 10 mL H2O. To prepare 50 mg/mL BCIP p-toluidine salt, dissolve 0.5 g in 10 mL DMF.
Is BCIP light sensitive?
BCIP/NBT -Alkaline Phosphatase Substrate Instructions for Use: Do Not Dilute. This solution is provided “ready-to-use” and may not perform correctly if diluted. The substrate is light sensitive and must be protected from light during incubation with the alkaline phosphatase-labeled probe.
What is Pnpp substrate?
PNPP (p-Nitrophenyl Phosphate, Disodium Salt) is a widely used substrate for detecting alkaline phosphatase in ELISA applications. When alkaline phosphatase and PNPP are reacted, a yellow water-soluble reaction product is formed.
What is an ELISA substrate?
Colorimetric substrates are commonly used in ELISA. The substrate is a crucial component for a successful ELISA assay. The choice of ELISA substrate depends on the enzyme used in the preceding detection step. Alkaline phosphatase (ALP) and horseradish peroxidase (HRP) require different substrates.
What is AP ELISA?
Alkaline phosphatase (AP) conjugates are commonly used as secondary detection reagents in ELISA, immunohistochemistry and western blot. See alkaline phosphatase AP-conjugated secondary antibodies with different specificities, including fragment and pre-adsorbed antibodies.
What does alkaline phosphatase do to Pnpp?
PNPP (p-Nitrophenyl Phosphate, Disodium Salt) is a widely used substrate for detecting alkaline phosphatase in ELISA applications. When alkaline phosphatase and PNPP are reacted, a yellow water-soluble reaction product is formed. This reaction product absorbs light at 405nm.
Which enzyme is used in ELISA?
The most commonly used enzyme labels are horseradish peroxidase (HRP) and alkaline phosphatase (AP). Other enzymes have been used as well; these include β-galactosidase, acetylcholinesterase, and catalase. A large selection of substrates is available commercially for performing ELISA with an HRP or AP conjugate.
What is the most common alkaline phosphatase substrate?
Bromochloroindoyl phosphate/nitro blue tetrazolium (BCIP/NBT) is the most commonly used of the chromogenic substrates for alkaline phosphatase. It is somewhat more sensitive than NABP/New Fuchsin but gives less contrast during microscopic observation.
How to use NABP-NF for the detection of Naphthol AS-TR phosphate?
Reagents for detection using NABP-NF (Steps 1–10) 1. Dissolve 1 mg of New Fuchsin in 0.25 mL of 2 n HCl. 2. Dissolve 1 mg of sodium nitrate in 0.25 mL of H 2 O. 3. Dissolve 10 mg of naphthol AS-TR phosphate in 0.2 mL of dimethylformamide. 4. Mix the New Fuchsin solution with the sodium nitrate solution. Shake for 1 min.
What is NABP/NABP/new fuchsin?
Naphthol-AS-BI-phosphate (NABP)/New Fuchsin produces an intense red product that is insoluble in alcohols as well as aqueous solutions and is compatible with a range of histochemical stains. Bromochloroindoyl phosphate/nitro blue tetrazolium (BCIP/NBT) is the most commonly used of the chromogenic substrates for alkaline phosphatase.
