| Progesterone is a gonadal hormone and an accurate indicator for ovarian activity.So monitoring progesterone concentration in milk is an effective method to prevent and treat reproductive disorders of dams. Detection of progesterone should be carried out according to the method that is efficient, sensitive and specific. The study prepared progesterone complete antigen using carbodiimide method, accordingly built up the indirect competitive ELISA progesterone test method in milk, and subsequently was converted to screen-printed electrode for rapid detection of progesterone in milk.Progesterone derivatives 11α-hydroxyprogesterone hemisuccinate was used as hapten which is with OVA to prepare complete antigen (P4-OVA) by the effect of coupling agent DDC;P4-OVA coupling proved successful by immunoassay and applied the relevant ELISA methods.We set up indirect competitive ELISA method for rapid detection of progesterone by prepared conjugate, and the corresponding conditions were optimized(e.g.coating concentration, monoclonal antibody concentration,hrp-antibody concentration). Detection range:0.21ng/mL-40ng/mL.This method is used to develop progesterone biosensor for determination in milk.It applied indirect competitive ELISA method to disposable screen-printed electrode, and determined the electric current changes of TMB-H2O2 substrate reaction under horseradish peroxidase by chronoamperometry.The peak currents of the biosensors linearly depended on the concentration of progesterone in the range of 0.16ng/mL-50ng/mL, with the limits of detection 0.16ng/mL.The selectivity of biosensor was satisfactory and there were no cross reactivities in the determinations for estradiol, estriol and testosterone. The relative standard deviation was less than 9%. recovery 95.8%-115.7%, the method was accutate and reliable. The biosensor can be conserved for 4 days at 4℃.Generally it showed that is well overall performance for the detection of progesterone in milk.
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