血红蛋白晚期糖基化终末产物的酶联免疫吸附分析法检测

Measurement of hemoglobin-advanced glycosylation end products b y competitive ELISA

采用体外孵育形成的RNase-AGEs免疫大白兔，制备抗RNase-AGEs抗血清，建立检测体内Hb-AGEs的竞争性酶联免疫吸附分析法(ELISA)。抗血清鉴定结果显示: 滴度为1:5000，亲和常数为6.33×106L/mol，能特异性结合Hb-AGEs上的AGEs抗原决定簇，灵敏度为0.8 AGE.U/mg Hb，批内、批间CV为4.9%和8.8%。2型糖尿病患者Hb-AGEs含量比正常对照者显著增高（P=0.0081），平均增高70.8%，与HbA1c呈正相关（r=0.368，P＜0.05)。表明该抗血清适合于体内AGEs的检测, Hb-AGEs可作为体内AGEs的一个循环标志物，将有助于糖尿病并发症的研究和防治。

In the present study, ribonuclease-AGE (RNase-AG Es) prepared in vitro was used as immunogen to immunize rabbits to prepare p olyclonal antiserum to AGE epitopes, and then, competitive enzyme-linked immunosorbent assay(ELISA) was d eveloped. The antiserum was found to react with RNase-AGEs, RNase, hemoglobin - AGEs(Hb-AGEs) and bovine serum albumin-AGE(BSA-AGE), but not with BSA and Hb . It had a titre of 1:5000 and a Ka of 6.33×106L/mol. The minimum detec tabl e amount was calculated to be 0.8 AGE.U/mg Hb for Hb-AGEs. CV within or a mon g batches was 4.9% or 8.8%, respectively. It was found that the Hb-AGEs le vel s in 50 patients with type 2 diabetes were 70.8% higher than those in 44 normal individuals( P ＜0.01), and were significantly positive correlation with he moglobin A 1C(HbA1C) in the patients(r=0.368, P ＜0.05). It was suggested t hat the a ntiserum proved suitable for the measurement of AGEs which are formed in vivo . Hb-AGEs is a circulating marker of in vivo -formed AGEs, whic h is helpful to investigation and treatment of diabetic complications.