General 25-Hydroxyvitamin D3 ELISA Kit

From Antibodies.com, this competitive Enzyme-Linked Immunosorbent Assay (cELISA) is designed for the in vitro quantitative determination of 25-Hydroxyvitamin D3 in serum, plasma or other biological fluids, with universal reactivity. The assay uses a pre-coated microplate format comprising 12 x 8 well strips, with absorbance measured on a colorimetric microplate reader at 450 nm.

The kit employs a competitive enzyme immunoassay technique in which 25-Hydroxyvitamin D3 in samples or standards competes with immobilized antigen for binding to a Biotinylated Anti-25-Hydroxyvitamin D3 antibody. Following incubation and washing, an HRP-Avidin conjugate is added to the wells, and a TMB substrate solution is then used to visualize the enzymatic reaction. The resulting blue product changes to yellow upon addition of an acidic stop solution, and the absorbance at 450 nm is inversely proportional to the amount of 25-Hydroxyvitamin D3 present.

According to the manufacturer's specification, the assay range is 3.12–200 ng/ml with a sensitivity of <0.83 ng/ml, and a total assay time of approximately 2 hours 30 minutes. Reported mean recoveries in undiluted samples are 91% for serum (range 80%–102%, n=5), 90% for EDTA plasma (range 81%–100%, n=5), and 84% for heparin plasma (range 80%–89%, n=5). Kit components are stored at +4°C or −20°C as instructed, and the kit should not be used past its expiration date.