Banu N, Hara H, Kataoka S, Egusa G, Yamakido M
Second Department of Internal Medicine, Hiroshima University School of Medicine, Japan.
Ann Clin Biochem. 1994 Sep;31 ( Pt 5):485-91. doi: 10.1177/000456329403100511.
We investigated the efficacy of polyethylene glycol (PEG) for effective and reproducible concentration of urinary type IV collagen prior to measurement by enzyme immunoassay (EIA). Human placental type IV collagen at low concentrations (5 and 10 micrograms/L) and urinary type IV collagen were readily precipitated by PEG-4000 added at a concentration of about 150 g/L in the presence of 0.5 g/L gamma-globulin. Type IV collagen measurement by EIA from PEG-concentrated urine samples showed complete recovery and good reproducibility. Analysis of size distribution by Sephacryl S-300HR gel chromatography and Western blotting following polyacrylamide gel electrophoresis confirmed that type IV collagen in PEG-concentrated urine samples was of high molecular weight comparable to that of human placental type IV collagen. After PEG concentration, type IV collagen was detectable by EIA even in the urine of healthy subjects. Significantly higher concentrations of urinary type IV collagen were found in 30 diabetic patients with nephropathy than in 20 healthy subjects [99.5 (8.9) micrograms/L, mean (SEM) versus 21.4 (2.6) micrograms/L, P < 0.0001]. Thus, urinary type IV collagen can be measured effectively by EIA following concentration with PEG. This method has potential for the assessment of the progression of diabetic nephropathy.
我们研究了聚乙二醇（PEG）在通过酶免疫测定（EIA）进行测量之前有效且可重复地浓缩尿Ⅳ型胶原的效果。在0.5 g/Lγ球蛋白存在的情况下，以约150 g/L的浓度添加PEG - 4000可使低浓度（5和10微克/升）的人胎盘Ⅳ型胶原和尿Ⅳ型胶原容易沉淀。通过EIA对PEG浓缩尿样进行Ⅳ型胶原测量显示完全回收且重复性良好。通过Sephacryl S - 300HR凝胶色谱分析大小分布以及聚丙烯酰胺凝胶电泳后的蛋白质印迹法证实，PEG浓缩尿样中的Ⅳ型胶原具有与人类胎盘Ⅳ型胶原相当的高分子量。PEG浓缩后，即使在健康受试者的尿液中，通过EIA也可检测到Ⅳ型胶原。在30例糖尿病肾病患者中发现尿Ⅳ型胶原浓度显著高于20例健康受试者[99.5（8.9）微克/升，均值（标准误）对21.4（2.6）微克/升，P < 0.0001]。因此，尿Ⅳ型胶原可通过PEG浓缩后用EIA有效测量。该方法在评估糖尿病肾病进展方面具有潜力。
