Immunoassay for diagnosis of Babesia infection

描述(由申请人提供)：该项目旨在开发一种用于临床诊断测试的巴贝斯虫抗体的酶免疫分析(EIA)。巴贝斯虫病是一种新出现的扁虱传播的寄生虫病，在免疫功能低下或其他方面虚弱的患者中可导致严重到致命的疾病，在流行区可在高达1%的人口中潜伏在血液中携带。最近发生的通过输血传播的巴贝斯虫病致死病例也使人们认识到这种病原体是一种重大威胁。 到血液供应。然而，目前还没有商业的、经过验证的和FDA批准的测试可以用于B.microti。巴贝斯虫病目前的诊断方法是免疫荧光染色、血涂片显微镜检查和/或聚合酶链式反应。这些程序都不容易适应常规的临床实验室使用。我们建议开发一种基于微孔板的酶联免疫吸附试验，使用免疫许可的新型人工合成的微小杆菌抗原，这些抗原以前已经被证明是诊断上有意义的标志。将在非流行区和其他对照地区的献血者血清中评估抗原的特异性。检测性能将在来自美国东北部和中西部流行地区的巴贝斯虫病患者的特征良好的血清上进行评估。为了确定可能进一步提高检测灵敏度的免疫优势表位，我们已经确定了一系列假定的巴贝斯虫抗原序列，这些序列将以重叠的多肽库形式与巴贝斯虫病患者血清进行筛选，这一策略已经导致发现了许多具有诊断价值的抗原。我们还提出了确定其他致病巴贝斯虫物种的免疫优势表位的策略，包括B.duncani/WA1和B.duncani/MO-1。通过这些策略发现的新的免疫优势表位将被合成为具有适当结构的多肽抗原，以整合到当前的分析格式中，这使得在单孔分析中结合多个不同的多肽成为可能。一旦开发出具有符合临床诊断要求的灵敏度和特异度的单孔分析格式，我们将为第二阶段的临床试验和FDA提交做准备。

DESCRIPTION (provided by applicant): This project is aimed at development of an enzyme immunoassay (EIA) for antibodies to Babesia for use as a clinical diagnostic test. Babesiosis is an emerging tick-borne parasitic disease which may cause severe to fatal illness in immunocompromised or otherwise weakened patients, and may be carried in the blood subclinically in up to 1% of the population in endemic areas. Recent cases of fatal transfusion-transmitted babesiosis have also led to the identification of this pathogen as a significant threat to the blood supply. However, currently no commercial, validated and FDA approved tests are available for B. microti. Babesiosis is currently diagnosed by immunofluorescence staining, microscopy of blood smears, and/or PCR. None of these procedures are easily adaptable to routine clinical laboratory use. We propose to develop a microplate-based ELISA using novel synthetic B. microti antigens licensed to Immunetics that have previously been shown to be diagnostically significant markers. Specificity of the antigens will be assessed in blood donor sera from nonendemic areas and other controls. Assay performance will be evaluated on well-characterized sera from babesiosis patients from both U.S. Northeast and Midwest endemic regions. To identify immunodominant epitopes that may further improve assay sensitivity, we have identified a series of putative Babesia antigenic sequences that will be screened with babesiosis patient serum in an overlapping peptide library format, a strategy that has led to discovery of a number of antigens of diagnostic value. We also propose strategies to identify immunodominant epitopes of other pathogenic Babesia species, B. divergens/MO-1 and B. duncani/WA1. Novel immunodominant epitopes discovered through these strategies will be synthesized as peptide antigens with an appropriate structure for incorporation into current assay formats, which enable combination of multiple distinct peptides in a single-well assay. Once a single-well assay format is developed with sensitivity and specificity characteristics consistent with clinical diagnostic requirements, we will prepare for clinical trials and FDA submission in Phase II.