Shi H, Ma Y, Humphrey J H, Craft N E
Division of Science, Northeast Missouri State University, Kirksville 63501, USA.
J Chromatogr B Biomed Appl. 1995 Mar 10;665(1):89-96. doi: 10.1016/0378-4347(94)00520-f.
We have developed a high-performance capillary electrophoresis (HPCE) method to analyze the retinol (vitamin A) concentration as retinol-retinol binding protein (holo-RBP) from microvolumes of serum (5-10 microliters) or one to two drops (approximately 20 microliters) of blood collected and air-dried on blood collection filter paper. A 0.64-cm diameter disk was cut from the dried whole blood specimens and the samples were dissolved in a pretreatment buffer and filtered. Filtrate was injected onto the HPCE column for analysis. The separation was carried out in a 60 cm x 50 microns I.D. fused-silica capillary and the running voltage was 20 kV. A He-Cd laser with a wavelength of 325 nm was used for excitation, and the fluorescence of the holo-RBP complex was monitored at 465 nm by a photodiode. A virtual linear relationship was obtained for the retinol concentrations between HPCE and HPLC for 28 serum samples, 19 dried venous blood samples and 9 capillary dried blood spot samples, indicating that valid measures of serum retinol can be obtained from one to two drops of capillary blood collected on filter paper. The absolute detection limit for retinol by HPCE is below 3 micrograms/l. The method is very useful for vitamin A level screening, especially for children and premature new-born babies.
我们开发了一种高效毛细管电泳（HPCE）方法，用于分析从微量血清（5 - 10微升）或采集并在采血滤纸上风干的一到两滴血（约20微升）中提取的视黄醇（维生素A）浓度，即视黄醇 - 视黄醇结合蛋白（全RBP）。从干燥的全血标本上切下直径为0.64厘米的圆盘，将样品溶解在预处理缓冲液中并过滤。滤液注入HPCE柱进行分析。分离在一根60厘米×50微米内径的熔融石英毛细管中进行，运行电压为20 kV。使用波长为325纳米的氦镉激光进行激发，通过光电二极管在465纳米处监测全RBP复合物的荧光。对于28个血清样本、19个干燥静脉血样本和9个毛细血管干血斑样本，HPCE与HPLC之间的视黄醇浓度呈现虚拟线性关系，这表明从采集在滤纸上的一到两滴血中可以获得有效的血清视黄醇测量值。HPCE对视黄醇的绝对检测限低于3微克/升。该方法对于维生素A水平筛查非常有用，特别是对于儿童和早产新生儿。
