异硫氰酸荧光素标记尿激酶的制备与性能研究

用异硫氰酸荧光素(FITC)对溶栓蛋白药物尿激酶(uPA)进行了荧光标记, 制备了荧光素标记的尿激酶(FITC-uPA), 经核磁谱及红外谱表征证实了反应能有效进行, 其荧光素/蛋白值(F/P)值为0.45. 尿激酶经荧光素标记后, 具有良好的荧光性质, 其荧光检测下限低达10-6 g?mL-1. 在1~100 ?g?mL-1范围内, 荧光强度与溶液浓度具有很好的线性关系, 可用于微/痕量蛋白质的定量/定性检测和跟踪标记. 荧光素标记后的尿激酶, 其流体力学直径与未标记的尿激酶基本一致, 其体外溶栓能力也与未标记的尿激酶相当, 说明FITC标记基本不影响尿激酶的生物活性, 是一种简单、有效的溶栓药物标记方法.

Preparation and Property of Fluorescein Labeled Urokinase

Fluorescein labeled urokinase (FITC-uPA) is synthesized by the reaction of fluorescein isothiocyanate (FITC) with urokinase (uPA), a kind of protein drug for thrombolysis. The reaction is confirmed by 1HNMR and FT-IR. The ratio (F/P) of fluorescein over protein is 0.45. The urokinase is found to possess good fluorescent property after fluorescein is labeled, and the detection limit of fluorescence is as low as 10-6 g?mL-1. There is obvious linear relationship between the fluorescein intensity and concentration of FITC-uPA within the concentration range of 1-100 μg?mL-1, which can be used for qualitative/quantitative detection and tracking of micro/trace proteins. FITC-uPA has similar hydrodynamic diameter and thrombolysis efficiency with the unlabeled urokinase. Based on the study, it can be concluded that the fluorescein-label is a simple and effective method for thrombolysis drug labeling without affecting the biological active of urokinase.