【摘要】：目的:建立測定人血漿可溶性血管內(nèi)皮鈣粘蛋白(sVE-cadherin)濃度的雙抗體夾心酶聯(lián)免疫吸附試驗(ELISA)方法,并探討其臨床應用價值。方法:應用原核表達的人VE-cadherin重組蛋白作為抗原免疫BALB/c小鼠制備單克隆抗體,采用棋盤格滴定法篩選出最佳雙抗體組合,其中檢測抗體采用辣根過氧化物酶(HRP)標記,建立檢測人血漿sVE-cadherin雙抗體夾心ELISA方法,并進行方法學評價。同時采用該法檢測28名健康體檢者和60例腫瘤患者的血漿sVE-cadherin濃度。結(jié)果:經(jīng)篩選共獲得6株單克隆抗體,通過篩查配對,成功建立檢測人血漿可溶性VE-cadherin的雙抗體夾心ELISA方法。該方法的檢測限為24.7 pg/ml,批內(nèi)變異系數(shù)(CV)為4.1%-7.7%,批間CV為8.7%-10.8%,平均回收率為96.7%。用該法檢測28名健康體檢者血漿VE-cadherin濃度為262.1±11.75 pg/ml,而用該方法檢測24例白血病患者血漿VE-cadherin濃度為173.9±17.98 pg/ml,顯著低于正常人(P0.01),14例胃癌患者血漿VE-cadherin濃度為311.7±25.24 pg/ml(P0.05),11例肺癌患者血漿VEcadherin濃度為206.8±25.01 pg/ml,均低于正常人(P0.05),11例其他腫瘤(9例乳腺癌,1例膠質(zhì)瘤,1例肝癌)患者血漿VE-cadherin濃度為310.7±11.82 pg/ml(P0.05)。結(jié)論:建立的雙抗體夾心ELISA試劑盒有較好的靈敏度及特異性,可用于人血漿中可溶性VE-cadherin含量的檢測,并可作為腫瘤診斷的一種新檢測指標。
[Abstract]:Aim: to establish a double antibody sandwich enzyme linked immunosorbent assay (ELISA) method for the determination of soluble vascular endothelial cadherin (sVE-cadherin) in human plasma and to explore its clinical value. Methods: Monoclonal antibodies were prepared by immunizing BALB/c mice with prokaryotic expressed human VE-cadherin recombinant protein. The best combination of double antibodies was screened by chessboard titration. The detection antibody was labeled with horseradish peroxidase (HRP). A method for the detection of double antibody sandwich ELISA (ELISA) in human plasma sVE-cadherin was established and evaluated. Plasma sVE-cadherin levels were measured in 28 healthy persons and 60 tumor patients. Results: six monoclonal antibodies were obtained and a double antibody sandwich ELISA method was successfully established for the detection of soluble VE-cadherin in human plasma by screening and pairing. The detection limit of this method was 24.7 pg/ml, within-run variation coefficient (CV) of 4.1-7.7.The CV between batches was 8.7- 10.8.The average recovery was 96.7. The plasma VE-cadherin concentration was 262.1 鹵11.75 pg/ml, in 28 healthy controls and 173.9 鹵17.98 pg/ml, in 24 leukemia patients (P0.01). The plasma VE-cadherin concentration of 14 patients with gastric cancer was 311.7 鹵25.24 pg/ml (P0.05), the plasma VEcadherin concentration of 11 patients with lung cancer was 206.8 鹵25.01 pg/ml, lower than that of normal controls (P0.05), 11 patients with other tumors (9 patients with breast cancer), 11 patients with other tumors (9 patients with breast cancer, 9 patients with breast cancer), The plasma VE-cadherin concentration in 1 case of glioma and 1 case of liver cancer was 310.7 鹵11.82 pg/ml (P0.05). Conclusion: the double antibody sandwich ELISA kit has good sensitivity and specificity. It can be used for the detection of soluble VE-cadherin in human plasma, and can be used as a new detection index for tumor diagnosis.
【作者單位】： 蘇州大學附屬第一醫(yī)院檢驗科;江蘇省血液研究所;蘇州大學唐仲英血液學研究中心;
【基金】：江蘇高校優(yōu)勢學科建設(shè)工程資助項目(YX21100214) 蘇州市第十五批科技發(fā)展項目(SYS201409)
【分類號】：R446.6

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