Liu Jia-Ming, Lu Qiao-mei, Wang Yan, Xu Shan-shan, Lin Xiao-mei, Li Long-Di, Lin Shao-Qin
Department of Chemistry, Zhangzhou Normal College, PR China.
J Immunol Methods. 2005 Dec 20;307(1-2):34-40. doi: 10.1016/j.jim.2005.09.007. Epub 2005 Oct 19.
Luminescent 20-nm silicon dioxide nanoparticles containing dibromofluorescein (D; particles denoted D-SiO(2)) were synthesized by the sol-gel method. In the presence of Pb(Ac)(2) as a heavy atom perturber, the particle can emit the intense and stable room temperature phosphorescence(RTP) signal on a polyamide membrane at the wavelength of lambdaex(max)/lambdaem(max) = 460/623 nm for D. Our research indicates that the specific immune reaction between goat-anti-human IgG antibody labeled with D-SiO(2) and human IgG can be carried out quantitatively on a polyamide membrane, and the phosphorescence intensity was evidently enhanced after the immunoreaction. Thus a new solid substrate-room temperature phosphorescence immunoassay (SS-RTP-IA) for determination of human IgG was established. The linear range of this method is 0.0624-20.0 pg spot(-1) of human IgG (corresponding to a concentration range of 0.156-50.0 ng ml(-1), sample volume: 0.40 microl spot(-1)). The regression equation of the working curve is DeltaIp = 94.39 + 17.00 m IgG (pg spot(-1)) (460/623 nm, r = 0.9998). Detection limit calculated as 3 Sb/k is 0.015 pg spot(-1). After elevenfold replicate measurement, RSD are 3.2% and 2.4% for samples containing 0.156 and 50.0 ng l(-1) IgG, respectively. This method is sensitive, accurate, and of high precision. And it has been applied to the determination of IgG in human serum with satisfactory results. Meanwhile, the mechanism of SS-RTP-IA based on an antibody labeled with nanoparticles containing dibromofluorescein luminescent molecules was discussed.
采用溶胶-凝胶法合成了含有二溴荧光素（D；标记为D-SiO₂的颗粒）的发光20纳米二氧化硅纳米颗粒。在作为重原子微扰剂的Pb(Ac)₂存在下，该颗粒可在聚酰胺膜上于波长λex(max)/λem(max)=460/623 nm（D的波长）处发射强烈且稳定的室温磷光（RTP）信号。我们的研究表明，用D-SiO₂标记的山羊抗人IgG抗体与人类IgG之间的特异性免疫反应可在聚酰胺膜上进行定量，免疫反应后磷光强度明显增强。由此建立了一种用于测定人IgG的新型固体基质-室温磷光免疫分析法（SS-RTP-IA）。该方法的线性范围为人IgG的0.0624 - 20.0 pg spot⁻¹（对应浓度范围为0.156 - 50.0 ng ml⁻¹，样品体积：0.40 μl spot⁻¹）。工作曲线的回归方程为ΔIp = 94.39 + 17.00 m IgG（pg spot⁻¹）（460/623 nm，r = 0.9998）。以3Sb/k计算的检测限为0.015 pg spot⁻¹。经过11次重复测量，对于含有0.156和50.0 ng l⁻¹ IgG的样品，相对标准偏差分别为3.2%和2.4%。该方法灵敏、准确且精密度高。并且已应用于人体血清中IgG的测定，结果令人满意。同时，还讨论了基于含有二溴荧光素发光分子的纳米颗粒标记抗体的SS-RTP-IA的机制。
