Song Eun-Young, Kim Kyung-Sook, Kim Kyoung-A, Kim Yung-Dai, Kwon Dur-Han, Byun Si-Myung, Kim Hee-Jung, Chung Tae-Wook, Choe Yong-Kyung, Chung Tai-Wha, Kim Cheorl-Ho
Cell Biology Laboratory, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-600, Korea.
Glycoconj J. 2002 Jul;19(6):415-21. doi: 10.1023/B:GLYC.0000004013.36690.78.
The glycoprotein UDP-N-acetylglucosamine: beta-D-mannoside-1,4-N-acetylglucosaminyltransferase-III (GnT-III) catalyzes the addition of N-acetylglucosamine via a beta-1, 4-linkage to the beta-linked mannose of the trimannosyl core of N-linked glycans. It has been reported that the expression of GnT-III increases in many oncogenically transformed cells and human hepatocellular carcinoma (HCC) tissues, and GnT-III enzyme activity in serum can be used for the detection and monitoring of primary hepatomas and hepatocellular carcinomas. A solid-phase enzyme-linked immunosorbent sandwich assay in which a polyclonal antibody (PAb) to aglycosylrecombinant GnT-III (AGR-GnT-III) and a monoclonal antibody (mAb) are employed as a capture protein and probe protein, respectively, is described. The sensitivity of the PAb-mAb sandwich assay, as determined by the dose-response effect for AGR-GnT-III, was 10 ng/ml. This assay was specific for GnT-III and did not detect beta-1, 6-N-acetylglucosaminyltrasferase-V (GnT-V). AGR-GnT-III concentrations in 377 serum specimens were determined by the PAb-mAb sandwich assay and the results were analyzed based on the disease category, using 1.99 microg/mL (AGR-GnT-III) as a cut-off value. The AGR-GnT-III level of 61 normal serum samples was 0.57 +/- 0.71 microg/ml (mean +/- SD). The results revealed an elevation in serum AGR-GnT-III levels in 60 of 86 patients (3.03 +/- 2.04 microg/ml) with liver cirrhosis (LC) and 86 of 91 patients (2.73 +/- 0.59 microg/ml) with chronic hepatitis (CH). By contrast, 3 of 61 normal subjects, 9 of 34 patients (1.02 +/- 1.03 microg/ml) with acute hepatitis and 8 of 38 patients (1.79 +/- 0.56 microg/ml) with a variety of non-hepatic diseases exhibited a slight increase above the cut-off value. These results indicate that serum AGR-GnT-III levels are elevated predominantly in LC or CH cases. Serum AGR-GnT-III concentration, as measured by the developed PAb-mAb sandwich assay, may be a useful differential marker as a diagnostic aid for CH and/or LC and warrants further investigations with expanded serum panels.
糖蛋白UDP-N-乙酰葡糖胺：β-D-甘露糖苷-1,4-N-乙酰葡糖胺基转移酶III（GnT-III）催化通过β-1,4-连接将N-乙酰葡糖胺添加到N-连接聚糖三甘露糖核心的β-连接甘露糖上。据报道，GnT-III在许多致癌转化细胞和人类肝细胞癌（HCC）组织中的表达增加，血清中的GnT-III酶活性可用于原发性肝癌和肝细胞癌的检测与监测。本文描述了一种固相酶联免疫吸附夹心测定法，其中分别使用针对无糖基重组GnT-III（AGR-GnT-III）的多克隆抗体（PAb）和单克隆抗体（mAb）作为捕获蛋白和探针蛋白。通过AGR-GnT-III的剂量反应效应测定，PAb-mAb夹心测定法的灵敏度为10 ng/ml。该测定法对GnT-III具有特异性，未检测到β-1,6-N-乙酰葡糖胺基转移酶-V（GnT-V）。采用PAb-mAb夹心测定法测定了377份血清标本中的AGR-GnT-III浓度，并以1.99 μg/mL（AGR-GnT-III）为临界值，根据疾病类别对结果进行了分析。61份正常血清样本的AGR-GnT-III水平为0.57±0.71 μg/ml（平均值±标准差）。结果显示，86例肝硬化（LC）患者中有60例（3.03±2.04 μg/ml）血清AGR-GnT-III水平升高，91例慢性肝炎（CH）患者中有86例（2.73±0.59 μg/ml）升高。相比之下，61例正常受试者中有3例、34例急性肝炎患者中有9例（1.02±1.03 μg/ml）以及38例各种非肝脏疾病患者中有8例（1.79±0.56 μg/ml）的AGR-GnT-III水平略高于临界值。这些结果表明，血清AGR-GnT-III水平主要在LC或CH病例中升高。通过所建立的PAb-mAb夹心测定法测得的血清AGR-GnT-III浓度，可能作为CH和/或LC诊断辅助的有用鉴别标志物，值得对更多血清样本进行进一步研究。
