聚乙二醇增敏亚甲蓝-四苯硼化钠离子缔合物荧光探针荧光猝灭法测定痕量蛋白质

在KH2PO4- Na2HPO4缓冲溶液中，离子缔合物[MB]+·[B(C6H5)4]–可发射强而稳定的荧光，牛血清蛋白（BSA）能使[MB]+·[B(C6H5)4]–的荧光信号显著猝灭，聚乙二醇(PEG)对荧光信号猝灭的有强的增敏作用，加PEG比不加PEG时，ΔF（= F0－F，其中，F0与F分别为试剂空白和试液的荧光强度）值提高了9.1倍，且ΔF与BSA含量具有良好的线性关系，据此建立了新型荧光探针荧光猝灭法测定痕量蛋白质的新方法。本方法的线性范围为0.11 ~ 88.0 ag/mL，检出限：22.0 ag /mL BSA，灵敏度很高，并成功用于人血清样品中蛋白含量的测定。同时探讨了新方法的反应机理。在相同条件下，新方法可分别测定BSA、人血清白蛋白(human serum albumin，HAS)、卵蛋白(ovalbumin，OVA )、γ-球蛋白(γ-globulin，γ-G)及血清、脑脊液样品中蛋白质总量。

Determination of Trace Protein by Methylene Blue‐Tetraphenylborate Fluorescence Probe Using Polyethylene Glycol as Sensitizer

An ionic association complex of methylene blue‐tetraphenylborate ([MB]⁺·[B(C₆H₅)₄]⁻) can emit a strong and stable fluorescence in KH₂PO₄‐Na₂HPO₄ buffer solution. In the presence of bovine serum albumin (BSA), the fluorescence signal of [MB]⁺·[B(C₆H₅)₄]⁻ can be sharply quenched, which can be further quenched by using polyethylene glycol (PEG) as a sensitizer where the ΔF (ΔF=F₀−F, F₀ and F are the fluorescence intensities of the blank reagent and the test solution, respectively) of the system with PEG is 9.1 times higher than that without PEG, showing PEG has strong sensitizing effect on the quenching of a fluorescence signal. And there is a good linear correlation between ΔF and the content of BSA. Thus, a new fluorescence probe for the determination of trace protein has been established, with the linear range of 0.11–88.0 pg·L⁻¹ and the detection limit of 22.0 ag·mL⁻¹ BSA. This sensitive method has been applied to the determination of protein in human serum samples with satisfactory results. And the reaction mechanism was also discussed. Under the same condition, the new method can be used to determie not only BSA, human serum albumin (HAS), ovalbumin (OVA) and γ‐globulin (γ‐G), respectively, but also the total protein in serum, brain and spinal cord.