Ultra-sensitive method for HIV p24 antigen detection

摘要 该项目的最终目标是开发一种商业照明设备， 系统试剂盒和原始ELISA诊断试剂盒用于检测人血清或血浆中的HIV p24抗原 光化学扩增法（PAM）在第一阶段和第二阶段开发。艾滋病病毒的鉴定方法 感染包括HIV抗体测试、通过病毒培养分离病毒、检测病毒RNA核酸测试 检测病毒p24抗原的试验和检测病毒酶逆转录酶的试验。这些方法 高度特异性，阳性结果几乎总是确认感染。然而，每一种都有其局限性， 使用必须适合于适当的测试情况。病毒核酸检测已经引入，但需要 先进的技术，敬业和训练有素的人员，而且非常昂贵。检测p24 抗原检测比核酸检测更简单，并为监测艾滋病毒感染提供了替代方法。的 p24抗原检测在血液筛查中也有价值，可用于急性感染的鉴别， 新生儿感染的诊断，以及检测已感染的培养物上清液中的抗原 用疑似感染病人的细胞接种。 目前，p24抗原检测相对不敏感，只能检测5-20 pg/ml。这个量 即使病毒正在活跃地复制，感染个体的血清中也可能不存在抗原。 因此，开发和使用更敏感的p24抗原检测方法具有很大的现实意义。正是本着这种 在第一阶段和第二阶段，我们开发了高度敏感的p24抗原检测， RNA病毒载量测试，适用于发达国家和发展中国家， 由于基础设施和实验室能力有限，无法进行核酸检测。 IIB期竞争性续期的具体目标是： 1.设计和制造一个预生产PAM照明装置。 2.设计并研制PAM检测系统试剂盒、HIV p24 ELISA试剂盒和第四代ELISA试剂盒 作为即用型ELISA + PAM诊断试剂盒用于HIV检测试剂盒。 3.根据FDA监管途径，使用500多个 临床样本。

Abstract The ultimate goal of the proposed project is to develop a commercial illumination device, detection system kit and original ELISA diagnostic kits for detection of HIV p24 antigen in human serum or plasma by a photochemical amplification method (PAM) developed within Phases I and II. The methods for identifying HIV infection include HIV antibody tests, viral isolation through viral culture, nucleic acid tests to detect viral RNA, tests to detect viral p24 antigen and tests to detect the viral enzyme reverse transcriptase. These methods are highly specific, and a positive result almost always confirms infection. However, each has limitations, and their use must be tailored to proper testing situations. Tests for viral nucleic acid have been introduced but require sophisticated technology, dedicated and well-trained personnel, and are very expensive. Tests to detect p24 antigen are simpler to perform than nucleic acid tests and offer alternatives for monitoring HIV infection. The p24 antigen test also can be of value in blood screening, for identification of acute infection, to assist in the diagnosis of infection in the newborn, and in detecting antigen in supernatants from cultures that have been inoculated with cells from a patient suspected of being infected. At present, p24 antigen tests are relatively insensitive, being able to detect only 5-20 pg/ml. This quantity of antigen may not be present in the serum of infected individuals, even when the virus is actively replicating. Therefore, development and the use of more sensitive p24 antigen tests has a great relevance. It is in this context that in Phases I and II we developed highly sensitive p24 antigen assays that are less expensive than RNA viral load tests and appropriate for use both in developed and developing countries in which the limitations of infrastructure and laboratory capability prohibit nucleic acid testing. The specific aims of Phase IIB Competing Renewal are: 1. To design and manufacture a pre-production PAM illumination device. 2. To design and develop a PAM detection system kit, HIV p24 ELISA kit and fourth generation ELISA kit for HIV detection as ready-to-use ELISA + PAM diagnostic kits. 3. To perform pre-clinical studies in accordance with FDA Regulatory pathway using over five hundred clinical samples.