应用蛋白质芯片检测多药耐药蛋白表达水平

Detection of multidrug-resistance proteins with protein array chips

目的探讨蛋白质芯片在检测白血病细胞多药耐药(MDR)蛋白表达中的价值。方法以人红白血病细胞系K562及其耐药细胞系K562/A02为实验研究对象。将3种耐药蛋白P糖蛋白(P-gP)、多药耐药相关蛋白(MRP1)和乳腺癌耐药蛋白(BCRP)相应的单克隆抗体固定在玻片上形成微阵列,细胞直接与固定在芯片上的耐药抗体阵列反应,电荷耦合器件(CCD)检测反应结果,并与流式细胞术测定的结果进行比较分析。结果在K562细胞中,蛋白质芯片检测到P-gP和BCRP表达率低,MRP1有较高水平的表达;在K562/A02细胞中,P-gP和MRP1均有高水平表达,BCRP表达率低。流式细胞术结果显示,K562细胞P-gP、MRP1和BCRP表达率分别为5.98%±2.19%、95.80%±3.98%和1.03%±0.45%;K562/A02细胞P-gP、MRP1和BCRP表达率分别为92.67%±1.80%、97.18%±1.02%和3.98%±0.37%。经统计学分析,两种方法结果一致(Ρ>0.05)。结论利用蛋白质芯片检测MDR蛋白结果可靠,具有高通量、低成本、制备简单、测定快速的优点。

Objective To evaluate the use of protein array chips in detection of muhidrug-resistance proteins. Methods Human crythrolcukcmic cell line K562 and its doxorubicin-resistant counterpart K562/ A02 were used in the study. Monoclonal antibodies against P-glycoprotein （ P-gP）, multidrug resistance- associated protein （MRP1） and breast cancer resistance protein （BCRP） were immobilized onto agarose film-coated glass. The antibody-cell binding was assessed by capturing K562 and K562/A02 cells. The protein array was observed under a microscope and the image was captured with a CCD camera. The expression levels of the three proteins were also measured by flow cytometry （ FCM ）. Results The expression of P-gP and BCRP in K562 was very low. However, MRP1 expression was high. P-gP and MRP1 were highly expressed in K562/A02, while the expression of BCRP was low. FCM results showed that the expression rate of P-gP, MRP1 and BCRP in K562 cells was 5.98% ±2.19% ,95.80% ± 3.98% , 1.03% ± 0.45%, respectively, while that in K562/A02 cells was 92.67% ± 1.80% ,97.18% ± 1.02% ,3.98% ± 0.37% , respectively. The results of protein array method are consistent with those of FCM （P 〉 0.05）. Conclusion h is feasible to develop a new protein array technique and to provide a novel method for multidrug resistant cell detection,with a high throughput, high specificity, simple procedure and low cost.