富勒醇-FITC磷光标记固体基质室温磷光法测定痕量碱性磷酸酶

开发了新的富勒醇、异硫氰酸荧光素和N,N-二甲基苯胺（F-ol-（FITC）n-DMA）标记试剂.基于F-ol的活性-OH与FITC游离的-COOH反应生成含有多个FITC分子的复合物（F-ol-（FITC）n-DMA）,增加WGA-AP-WGA-F-ol FITC-DMA（WGA和AP分别为麦胚凝集素与碱性磷酸酶）生物靶的发光分子数而进一步提高SSRTP（固体基质室温磷光法）的灵敏度.用F-ol-（FITC）n-DMA标记WGA方法的检出限为0.15 ag spot-1（F-ol）与0.097 ag spot-1（（FITC）,比F-ol-DMA、FITC-DEA单发光分子标记WGA方法的检出限（0.20 ag spot-1（F-ol）,0.22 ag spot-1（FITC）低.用于血清试样中AP含量的测定结果与酶联免疫法相吻合.

Solid Substrate RoomTemperature Phosphorimetry for the Determination of Trace Alkaline Phosphatase Using Fullerol-fluorescein Isothiocyanate as Labelling Reagent

A new phosphorescent labelling reagent consisting of fullerol,fluorescein isothiocyanate and N,N-dimethylaniline（F-ol-（FITC）n-DMA） was developed.The mode of action is based on the reactivity of the active-OH group in F-ol with the-COOH group of FITC to form an F-ol-（FITC）n-DMA complex containing several FITC molecules.F-ol-（FITC）n-DMA increased the number of luminescent molecules in the biological target of WGA-AP-WGA-F-ol-（FITC）n-DMA（WGA and AP are wheat germ agglutinin and alkaline phosphatase,respectively） which improved the sensitivity using solid substrate room temperature phosphorimetry（SSRTP） detection.The proposed method provided high sensitivity and strong specificity for WGA-AP.The limit of detection（LD） was 0.15 ag AP spot-1 for F-ol and 0.097 ag AP spot-1 for FITC in F-ol-（FITC）n-DMA,which was lower than the method using single luminescent molecules of F-ol-DMA and FITC-DMA to label WGA（0.20 ag AP spot-1 for F-ol-DMA and 0.22 ag AP spot-1 for FITC-DMA）.Results for the determination of AP in human serum were in good agreement with those obtained by enzyme-linked immunosorbent assay.The mechanism of F-ol-（FITC）n-DMA labelling of WGA was discussed.