Campbell A K, Patel A
Biochem J. 1983 Oct 15;216(1):185-94. doi: 10.1042/bj2160185.
A chemiluminescent derivative of cyclic AMP, aminobutylethylisoluminol succinyl cyclic AMP (ABEI-scAMP), was synthesized in order to develop a homogeneous immunoassay based on non-radiative energy transfer. ABEI-scAMP was chemiluminescent (5.1 X 10(18) luminescent counts X mol-1 at pH 13), pure (greater than 95%) stable and immunologically active. A conventional immunoassay was established using ABEI-scAMP and a solid-phase anti-(cyclic AMP) immunoglobulin G which could detect cyclic AMP at least down to 25fmol. A homogeneous immunoassay for cyclic AMP was established by measuring the shift in wavelength from 460 to 525nm which occurred when ABEI-scAMP was bound to fluorescein-labelled anti-(cyclic AMP) immunoglobulin G. The assay was at least as sensitive as the conventional radioimmunoassay using cyclic [3H]AMP and could measure cyclic AMP over the range 1-1000nM. The homogeneous chemiluminescent energy transfer assay was also able to quantify the association and dissociation of antibody-antigen complexes. Chemiluminescence energy transfer occurred between fluorescein-labelled antibodies and several other ABEI-labelled antigens (Mr values 314-150000) including progesterone, cyclic GMP, complement component C9 and immunoglobulin G. The results provide a homogeneous immunoassay capable of measuring free cyclic AMP under conditions likely to exist inside cells.
为了开发一种基于非辐射能量转移的均相免疫测定法，合成了环磷酸腺苷（cAMP）的化学发光衍生物，氨基丁基乙基异鲁米诺琥珀酰环磷酸腺苷（ABEI-scAMP）。ABEI-scAMP具有化学发光性（在pH 13时为5.1×10¹⁸发光计数×mol⁻¹），纯度高（大于95%），稳定性好且具有免疫活性。使用ABEI-scAMP和固相抗（环磷酸腺苷）免疫球蛋白G建立了一种传统免疫测定法，该方法能够检测低至25飞摩尔的环磷酸腺苷。通过测量ABEI-scAMP与荧光素标记的抗（环磷酸腺苷）免疫球蛋白G结合时发生的波长从460nm到525nm的变化，建立了一种环磷酸腺苷的均相免疫测定法。该测定法至少与使用环[³H]AMP的传统放射免疫测定法一样灵敏，并且能够在1-1000nM范围内测量环磷酸腺苷。这种均相化学发光能量转移测定法还能够定量抗体-抗原复合物的结合和解离。化学发光能量转移发生在荧光素标记的抗体与其他几种ABEI标记的抗原（分子量314-150000）之间，包括孕酮、环鸟苷酸、补体成分C9和免疫球蛋白G。这些结果提供了一种能够在细胞内可能存在的条件下测量游离环磷酸腺苷的均相免疫测定法。
