|Other Titles||Fluoroimmunoassay of estradiol based on the thermo-sensitive hydrogel|
An indirect competitive fluoroimmunoassay of estradiol was developed based on the thermo-sensitive hydrogel, poly-N-isopropylacrylamide(pNIPA). Antigen was conjugated,to pNIPA to form antigen bound thermally reversible hydrogel (pNIPA-Ag) after the antigen was synthesized by covalent bonding estradiol to bovine serum albumin. The anti-estradiol monoclonal antibody(McAb) was prepared by an in vivo method with McAb secreting hybridomas. Fluorescein isothiocyanate (FITC) labeled McAb (McAb-F) was prepared by. bonding FITC to purified McAb. Then free estradiol reacted with limited amount of McAb-F competing with pNIPA-Ag. Separation of the immune complex, pNIPA-Ag-McAb-F, was carried out by centrifuging and precipitating at a temperature above critical solution temperature of the hydrogel. After the above immune complex was dissolved, the fluorescence intensity was measured by a Hitachi M-850 fluorospectrometer. The calibration curve of free estradiol covered a range of 10-625 ng/mL, and the detection limit of the presented method for free estradiol was 3.2 ng/mL. The cross-reactions and the recoveries of the immunoassay were also investigated.
|Appears in Collections:||化学与分子工程学院|