Development of enzyme immunoassay for chromogranin A (CgA) and profiling of plasma CgA concentrations in the cow
Development of enzyme immunoassay for chromogranin A (CgA) and profiling of plasma CgA concentrations in the cow
复制标题
开发嗜铬粒蛋白 A (CgA) 酶联免疫分析方法并分析奶牛血浆 CgA 浓度
DOI：
10.1111/asj.13542
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Takahashi T.
中科院分区：
文献类型：
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作者：
Saito C;Ido-Matsumoto H;Kumazaki H;Kubo T;Kanazawa T;Takahashi T.
本研究的目的是建立并表征牛嗜铬粒蛋白 A (bCgA) 的同源免疫测定法，并分析妊娠早期血浆 bCgA 浓度。我们合成了对应于 bCgA 氨基酸序列 341-355 的寡肽用于免疫兔子，并合成了对应于 bCgA 氨基酸序列 336-365 的肽作为生物素化示踪剂和参考标准。重组 bCgA 蛋白也在大肠杆菌溶解液中产生。剂量依赖性位移曲线是从 1 至 1,000 nM 的参考标准中获得的。位移曲线显示合成肽的参考标准与系列稀释的血浆样品或本研究中产生的重组bCgA蛋白之间存在良好的关系。测定灵敏度定义为低于零标准的两个标准差的值，计算结果为 0.46 nM。测定内和测定间变异系数分别为6.48%和13.4%。非妊娠动物妊娠早期血浆 bCgA 浓度的变化。本研究的结果表明，测定血浆 bCgA 浓度可作为评估牛生理状态的措施。
The objectives of this study were to establish and characterize a homologous immunoassay for bovine chromogranin A (bCgA) and to profile plasma bCgA concentrations during early pregnancies. We synthesized oligopeptide corresponding to the amino acid sequence 341–355 of bCgA for immunizing rabbits and peptide corresponding to the amino acid sequence 336–365 of bCgA for both a biotinylated tracer and reference standards. Recombinant bCgA protein was also generated inEscherichia colilysate. Dose‐dependent displacement curves were obtained from 1 to 1,000 nM of the reference standards. The displacement curves showed a good relationship between the reference standards of the synthetic peptide and the serially diluted plasma sample or recombinant bCgA protein generated in the present study. The assay sensitivity defined as the value of two standard deviations below the zero standard was calculated as 0.46 nM. The intraassay and interassay coefficients of variation were 6.48% and 13.4%, respectively. Changes in the plasma bCgA concentrations in early pregnancies undulated in nonpregnant animals. The results of the present study suggest that assaying plasma bCgA concentrations could be utilized as measures to evaluate the physiological status of cattle.