Determination of Bt Protein in transgenic cotton seeds by double antibody sandwich enzyme-linked immunosorbent assay.

Bt Cry1Ac protein sandwich enzyme-linked immunosorbent assay was developed with anti-Bt Cry1Ac protein mouse monoclonal antibody(mAb) and rabbit polyclonal antibody produced by Chemical Regulation Research Center(CRRC) of China Agricultural University(CAU).The linear range of the method was 0.78-50.0 ng.mL-1.The linear equation was y = 0.6634x1.7387,and the determinative coefficient was 0.992.The specificity obtained with the established assay was confirmed and verified by the commercial Bt Cry1Ac protein kit and could be qualitatively and quantitatively used to detect the Bt Cry1Ac protein.By the assay,identification of hybrid F1and F2with either of the parents is non-transgenic Bt cotton showed that,all the seeds of F1were positive and the ratio of seeds positive to negative of F2was 3∶1.The assay was also used to imitate AP detection and the limit of AP was 1∶110 for single seed which contained more than 140 ng of Bt Cry1Ac protein.