基于磁珠的ABO血型蛋白质芯片的构建及初步应用分析

Construction and application of magnetic bead-based protein microarray for ABO blood typing

目的构建一种以磁珠为标记物的,用于血型检测的可视化蛋白质芯片。方法用人免疫球蛋白G(IgG)及磁珠标记的抗人IgG进行琼脂糖浓度、点样浓度、温度和固定时间等实验条件的摸索。根据优化的条件,经过铺片、点样、固定、封闭等程序构建ABO血型检测蛋白质微阵列,加入血液样品进行反应,最后用磁珠标记的抗体进行检测。结果通过条件的优化发现,在1.0%的琼脂糖基片表面,以100μg/mL的蛋白质稀释液点样,之后在24℃固定8h,能够得到更优的信号强度。检测不同稀释度的抗原抗体,正定型效价可达到1∶819 2,反定型可达到1∶4 096。检测14例临床样品,结果样点清晰、肉眼可见,且结果判定与试管法一致性达100%。结论成功构建了基于磁珠的血型检测可视化蛋白质微阵列。

Objective To construct a visual blood typing protein microarray based on magnetic bead .Methods The optimal experiment conditions ,such as agarose concentration ,temperature ,immobilization concentration and time ,were explored on glass-slide with human IgG and anti-human IgG antibody coated magnetic bead .Based on the optimal conditions ,an ABO blood typing protein microarray was established by coating ,printing ,immobilizing and blocking .Then the blood samples were added to the microarrays ,and detected with antibody-conjugated magnetic beads .Results When the class slide was coated with 1% agarose ,and spotted with the protein probes （100 mg/mL in printing buffer） on the fabricated substrates and incubated in a humid chamber at 24 ℃ for 8 h ,better results could be obtained .The forward test titer was up to 1∶819 2 ,and the reverse test was 1∶409 6 by detecting different dilu-tion rate of samples with the arrays .14 clinical specimens were tested with the arrays and tube methods ,and the re-sults were same .Each positive spot on the arrays was strong and visible easily to the naked eye .Conclusion A sys-tem of magnetic bead-based protein array for ABO blood group was successfully constructed .