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目的:评估重组融合蛋白谷胱甘肽S-转移酶(GST)-肌钙蛋白I(TnI)(28～110aa)的实用价值。方法:诱导表达菌株BL21-PGEX-4T-3/TnI(84～330bp)表达GST-TnI(28～110aa),谷胱甘肽Sepharose-4B亲合层析柱纯化融合蛋白,经自动免疫分析仪检测其免疫反应性。采用双抗体夹心ELISA法检测稳定性,并与天然提纯心肌肌钙蛋白I(cTnI)抗原进行比较。结果:GST-TnI(28～110aa)融合蛋白表达水平高,具较高免疫反应性。在37℃条件下,GST-TnI(28～110aa)融合蛋白第3天免疫反应性仍保留60%,而人心肌提纯的cTnI在第3天免疫反应性几近丧失。结论:GST-TnI(28～110aa)融合蛋白具较高的免疫反应性,与天然cTnI相比较,重组GST-TnI融合蛋白稳定性好,有望作为cTnI测定的候选参考标准品。
Objective: To evaluate the practical value of recombinant fusion protein glutathione S - transferase (GST) - troponin I (TnI) (28 ~ 110aa). Methods: The fusion protein was expressed by GST-TnI (28-110 aa) and glutathione Sepharose-4B affinity chromatography on BL21-PGEX-4T-3 / TnI (84-330 bp) The immunoreactivity was tested. The stability was measured by double antibody sandwich ELISA and compared with native purified cardiac troponin I (cTnI) antigen. Results: The expression of GST-TnI (28-110 aa) fusion protein was high, with high immunoreactivity. The immunoreactivity of GST-TnI (28-110 aa) fusion protein remained at 60% on day 3 at 37 ° C, whereas the loss of immunoreactivity was almost abolished on day 3 after cTnI purification in human myocardium. CONCLUSION: GST-TnI (28-110 aa) fusion protein has high immunoreactivity. Compared with natural cTnI, the recombinant GST-TnI fusion protein has good stability and is expected to be a candidate reference standard for cTnI assay.