Mahboudi Fereidoun, Irina Nickolaeva A, Chevalier Alexander, Ghadiri Ataallah, Adeli Ahmad, Amini-Bavil-Olyaee Samad, Barkhordari Farzaneh, Farzamfar Bardia, Alinejad Mahtab
Biotechnology Department, Pasteur Institute of Iran, Tehran, Iran.
J Biotechnol. 2006 Sep 1;125(2):295-303. doi: 10.1016/j.jbiotec.2006.03.021. Epub 2006 May 2.
The objective of this study was expression of a recombinant fusion protein p24-gp41 to gain a proper folding pattern of the proteins which could be recognized by specific antibodies against human immunodeficiency virus type 1 (HIV-1) for development of a reliable serodiagnostic kit. Serodiagnostic method using enzyme-linked immunosorbent assay (ELISA) with the expressed recombinant fusion protein p24-gp41 was carried out to test the sensitivity and specificity of the protein using human sera and various reference panels from Boston Biomedica Inc. (BBI). The level of the expression was determined to be 30% and the final recovery from fermentation and purification process was calculated as 80 mg/L with more than 98% purity. The developed ELISA assay was demonstrated to have 100 and 99.5% sensitivity and specificity, respectively, detecting anti-HIV-1 antibody using 900 positive and 10,000 negative human sera. The developed assay showed reliable results in comparison with other reference HIV ELISA kits using various BBI panels as well. In conclusion, the recombinant fusion protein p24-gp41 was expressed and used to develop a serodiagnostic kit for screening of the HIV-1 with high sensitivity (100%) and specificity (99.5%) which could be useful for screening large groups of blood donors.
本研究的目的是表达重组融合蛋白p24-gp41，以获得能被抗1型人类免疫缺陷病毒（HIV-1）特异性抗体识别的蛋白质正确折叠模式，从而开发一种可靠的血清学诊断试剂盒。采用酶联免疫吸附测定（ELISA）方法，利用表达的重组融合蛋白p24-gp41进行血清学诊断，以检测该蛋白对人血清和来自波士顿生物医学公司（BBI）的各种参考样本的敏感性和特异性。测定表达水平为30%，经发酵和纯化过程后的最终回收率计算为80 mg/L，纯度超过98%。所开发的ELISA检测方法在检测900份阳性和10000份阴性人血清中的抗HIV-1抗体时，敏感性和特异性分别为达到100%和99.5%。与使用BBI各种样本的其他参考HIV ELISA试剂盒相比，所开发的检测方法也显示出可靠的结果。总之，表达了重组融合蛋白p24-gp41，并用于开发一种血清学诊断试剂盒，用于筛查HIV-1，具有高敏感性（100%）和特异性（99.5%），可用于筛查大量献血者群体。
