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[目的]制备鼠抗人PCNA Mab与人IgG的交联物作为临床分析阳性参考品。[方法]利用杂交瘤技术制备鼠抗人PCNA Mab,在SPDP的作用下与人IgG交联,建立ELISA方法评估其作为参考品的可行性。[结果]获得2株鼠抗人PCNA杂交瘤细胞;免疫印迹表明鼠抗人PCNA Mab与人IgG交联后对人PCNA和抗人IgG抗体均具有反应性;以该交联物作为参考品建立的ELISA中,300μg/L~4.69μg/L范围内,R2达0.994、准确度为88.26%~99.66%、相对标准偏差4.66%~17.95%;稀释400倍后,血清基质对检测结果没有明显干扰;SLE病人样本(n=41)的ELISA均值和高值均大于健康人(n=22);临床免疫膜条确认的阴阳性样本的ELISA结果分别16.44±1.30μg/m L和6.05±0.50μg/m L,与判别结果相关。[结论]鼠抗人PCNA Mab-人IgG交联物可作为阳性参考品应用于PCNA自抗体的免疫分析。
[Objective] To prepare a cross-linked product of mouse anti-human PCNA Mab and human IgG as a positive reference for clinical analysis. [Method] The mouse anti-human PCNA Mab was prepared by hybridoma technology and cross-linked with human IgG under the action of SPDP. The feasibility of using ELISA to evaluate its use as reference material was established. [Result] The two anti-human PCNA hybridoma cells were obtained. Immunoblotting showed that the anti-human PCNA Mab was reactive with human PCNA and anti-human IgG antibody after cross-linking with human IgG. The cross-linked product was used as a reference , The R2 reached 0.994 with the accuracy of 88.26% -99.66% and the relative standard deviation of 4.66% -17.95% in the range of 300μg / L ~ 4.69μg / L. After diluted 400 times, the serum matrix did not significantly interfere with the test results (N = 41) were higher than that of healthy people (n = 22). ELISA results of negative and positive samples confirmed by clinical immunoprecipitation were 16.44 ± 1.30μg / m L and 6.05 ± 0.50μg / m L, and the results of the discrimination. [Conclusion] The mouse anti-human PCNA Mab-human IgG conjugate can be used as a positive reference for the immunoassay of PCNA autoantibodies.