Khosravi M J, Papanastasiou-Diamandi A
CyberFluor Inc., Toronto, Ontario, Canada.
Clin Chem. 1993 Feb;39(2):256-62.
We evaluated the effect of hapten heterology on free thyroxine (FT4) immunoassays involving the biotin-streptavidin system and time-resolved fluorometry. We compared protein derivatives of thyroxine (T4) and triiodothyronine (T3) as solid-phase antigen or biotinylated protein-tracer conjugate for competitive (or sequential) binding to a mouse anti-T4 monoclonal antibody. In both one- and two-step assays, the heterologous combination of the antibody and T3 conjugates showed superior standard curve sensitivity but up to eightfold lower zero standard signal (B(o)) when the same amounts of antibody and conjugates were used. The improved sensitivity was not altered when the amount of coupled T3 was increased to obtain a B(o) value similar to that of the homologous combination of antibody and T4 conjugates. In the two-step format, the sensitivity of the homologous assay was insufficient for routine use, consistent with displacement of bound T4 during the antibody back-titration step (demonstrated in the T4 displacement experiment with excess conjugate). Results from the one-step (labeled antibody) heterologous assay for approximately 85 clinical samples correlated well with those from an immunofluorometric assay and a two-step radioimmunoassay. The assay was not affected by a wide variation in endogenous serum concentrations of T4-binding globulin and albumin.
我们评估了半抗原异源性对涉及生物素 - 链霉亲和素系统和时间分辨荧光法的游离甲状腺素（FT4）免疫测定的影响。我们比较了甲状腺素（T4）和三碘甲状腺原氨酸（T3）的蛋白质衍生物作为固相抗原或生物素化蛋白质 - 示踪剂共轭物，用于与小鼠抗T4单克隆抗体进行竞争性（或顺序性）结合。在一步法和两步法测定中，当使用相同量的抗体和共轭物时，抗体与T3共轭物的异源组合显示出更好的标准曲线灵敏度，但零标准信号（B(o)）低至八倍。当增加偶联T3的量以获得与抗体和T4共轭物的同源组合相似的B(o)值时，提高的灵敏度并未改变。在两步法中，同源测定的灵敏度不足以用于常规使用，这与抗体反滴定步骤中结合的T4的置换一致（在过量共轭物的T4置换实验中得到证明）。一步法（标记抗体）异源测定对约85个临床样本的结果与免疫荧光测定和两步放射免疫测定的结果相关性良好。该测定不受血清中T4结合球蛋白和白蛋白内源性浓度广泛变化的影响。
