Development of a screen-printed carbon electrochemical immunosensor for picomolar concentrations of estradiol in human serum extracts.
Investigations into the development of a prototype electrochemical immunosensor for estradiol (E(2)) are described. After optimising reagent loadings in a 96-well enzyme-linked immunosorbent assay (ELISA), antibodies (rabbit anti-mouse IgG and monoclonal mouse anti-E(2)) were immobilised by passive adsorption onto the surface of screen-printed carbon electrodes (SPCEs). A competitive immunoassay was then performed using an alkaline-phosphatase (ALP)-labelled E(2) conjugate. Calibration plots for E(2) buffer standards, performed colorimetrically on the SPCEs using a para-nitrophenyl phosphate substrate solution, were in good agreement with ELISA calibration plots. Electrochemical measurements were then performed using differential pulse voltammetry (DPV) following the production of 1-naphthol from 1-naphthyl phosphate. The calibration plot of DPV peak current versus E(2) concentration showed a measurable range of 25-500 pg/ml with a detection limit of 50 pg/ml. A coefficient of variation of between 13.0 and 15.6% was obtained for repeat measurements. The immunosensor was applied to the determination of E(2) in spiked serum, following an extraction step with diethyl ether. A mean recovery for the method of 102.5% was obtained with a CV of 19.1%. The options available for further development of the sensor regarding precision, limit of detection and direct sample analysis are discussed.
Pemberton
R M
RM
Centre for Analytical, Materials and Sensors Science, University of the West of England, Bristol BS16 1QY, UK.
Mottram
T T
TT
Hart
J P
JP
D000911:Antibodies, Monoclonal / Q000737:chemistry; D019921:Biotinylation; D002138:Calibration; D002244:Carbon / Q000737:chemistry*; D003124:Colorimetry; D004563:Electrochemistry / Q000379:methods*; D004566:Electrodes; D004797:Enzyme-Linked Immunosorbent Assay; D004958:Estradiol / Q000097:blood*; D006801:Humans; D007118:Immunoassay; D007074:Immunoglobulin G / Q000737:chemistry; D008956:Models, Chemical; D009281:Naphthalenes / Q000737:chemistry; D009284:Naphthols / Q000737:chemistry; D009943:Organophosphorus Compounds / Q000737:chemistry
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D009281:Naphthalenes; D009284:Naphthols; D009943:Organophosphorus Compounds; C029350:1-naphthol; C014694:naphthyl phosphate; D004958:Estradiol; D002244:Carbon