Abstract：
Detecting progesterone (P4) concentration in cow serum is essential for monitoring the pregnancy progress after fertilization and is significant for the dairy farming industry and veterinary medicine. This study reports enzyme-free immunomagnetic beads (IMBs)-based competitive immunoassay for detecting P4 by P4-bovine serum albumin (BSA)-modified biosensors. The anti-P4 antibody-conjugated IMBs serve as collectors to capture P4 in undiluted serum samples to prevent the biosensor surface from biosample contamination and as insulated labels to report the electron-transfer resistance signal of electrochemical impedance spectroscopy (EIS) measurement. The IMBs and P4-containing samples were mixed for 15-30 min, capable of obtaining stable P4@IMB complexes. The 0.2-kGauss pulsed magnetic field (PMF) of the 20-s pulse width and 20-s relaxation time applied for 5 min can shorten the immunoreaction time between the P4@IMBs and the P4-BSA-modified biosensor and reduce the IMB\'s nonspecific adsorption on the biosensor surface. This competitive immunoassay\'s cut-off value and detection limit were 7.71 ng/mL and 7.33 ng/mL, respectively, which is lower than the serum\'s P4 plateau concentration (over 8 ng/mL) of dairy cows on days 6-16 of estrus cycles and that in pregnancy. The IMB-based immunoassay combining the PMF attraction and the label-free EIS measurement exhibits promising potential for rapidly detecting P4 in undiluted serum.
摘要：
检测奶牛血清中的孕酮（P4）浓度对于监测受精后的妊娠进展至关重要，对于奶牛养殖业和兽医学具有重要意义。这项研究报告了基于无酶免疫磁珠（IMB）的竞争性免疫测定法，用于通过P4-牛血清白蛋白（BSA）修饰的生物传感器检测P4。抗P4抗体缀合的IMB用作收集器以捕获未稀释的血清样品中的P4以防止生物传感器表面受到生物样品污染,并且用作绝缘标记以报告电化学阻抗谱(EIS)测量的电子转移电阻信号。将IMB和含P4的样品混合15-30分钟，能够获得稳定的P4@IMB复合物。脉冲宽度为20s，弛豫时间为5min的0.2-k高斯脉冲磁场（PMF）可以缩短P4@IMB与P4-BSA修饰的生物传感器之间的免疫反应时间，并减少IMB在生物传感器表面的非特异性吸附。这种竞争性免疫测定的临界值和检测限为7.71ng/mL和7.33ng/mL，分别,在发情周期的第6-16天和怀孕期间，低于奶牛的血清P4平台浓度（超过8ng/mL）。基于IMB的免疫测定法结合了PMF吸引力和无标记的EIS测量，具有快速检测未稀释血清中P4的潜力。
