Improved Chlamydia Test Using Aptamers on a Portable POC Microfluidic Platform

描述（由申请人提供）：本项目的基本目标是开发一种高保真度、基于适体的生殖器衣原体感染检测方法，可以快速定量地测定不同血清型沙眼衣原体的感染水平。最终目标是将这种检测方法集成到一种廉价的便携式微流体设备中，可以在任何医生的办公室中使用。这一装置将提供一些服务不足人口重要健康方面的重要信息。关键因素是一个血清型特异性DNA适体家族，在检测沙眼衣原体方面表现出非常高的灵敏度和特异性（99,999 %）。DNA适体作为抗体的增强替代品，并在三明治ELISA的微流控版本中与目标微生物特异性反应，该三明治ELISA是由Claros Diagnostics （Cambridge MA）团队成员之一最近开发的便携式手持设备实现的。提出了两阶段方案。在约翰霍普金斯大学（Baltimore， MD）的国际性病、呼吸和生物威胁研究实验室验证沙眼衣原体适体检测是第一阶段的高潮。该计划的第二阶段将侧重于将检测生物化学和仪器网格结合起来，在一个新的护理点（医生办公室）检测沙眼衣原体。提案小组积累的初步数据包括约翰霍普金斯大学Charlotte A. Gaydos教授小组带来的关于沙眼衣原体感染的临床和社会方面的广泛经验和深刻知识。Claros Diagnostics的微流体检测平台是一种小型设备，能够处理微型生物样品，并允许对微升液体体积进行快速重复性处理。这种新型核酸适体选择技术是由Infoscitex公司开发的，并已成功应用于生产高质量的DNA适体，用于美国国防部的应用，如检测炭疽毒素和传染性朊病毒。

DESCRIPTION (provided by applicant): The fundamental goal of this program is to develop a point of care high fidelity, aptamer based assay for genital Chlamydia infections that allows fast and quantitative determination of levels of infection with different sero-types of Chlamydia trachomatis. The ultimate goal is to have this assay be integrated into an inexpensive portable microfluidics device that can be used in any physician's office. This device will provide vital information on important health aspects of some underserved populations. The key element is a family of serotype specific DNA aptamers that exhibit very high sensitivity and specificity (99,999 %) of detection of C. trachomatis. The DNA aptamers act as an enhanced replacement for the antibodies and specifically react with the target microorganisms in a microfluidic version of sandwich ELISA realized in a portable hand held device recently developed by one of the team members, Claros Diagnostics (Cambridge MA). A two phase program is proposed. The Phase I culminates in the validation of the aptamer assay for Chlamydia trachomatis in the International STD, Respiratory, and Biothreat Research Laboratory of Johns Hopkins University (Baltimore, MD). The Phase II of the program will focus on bringing together the detection biochemistry and instrumental grid in a new point of care (physician's office) assay for Chlamydia trachomatis. The preliminary data accumulated by the proposal team encompass extensive experience and profound knowledge of clinical and societal aspects of C. trachomatis infections brought by the group of Professor Charlotte A. Gaydos, Johns Hopkins University. Claros Diagnostics' microfluidics detection platform realized as a small device is capable of dealing with microscale bio-samples and allows rapid reproducible handling of microliter liquid volumes. The novel aptamer selection technology has been developed by Infoscitex Corporation and has been successfully applied to produce high quality DNA aptamers for the US Department of Defense applications, such as detection of anthrax toxin and infectious prions. PUBLIC HEALTH RELEVANCE: Currently, the most sensitive diagnosis of genital chlamydiosis employs quantitative real-time polymerase chain reaction qRT-PCR. As recently reported, the major outer membrane protein (omp-1) -gene and a L- serovar-specific region of the polymorphic protein H (pmp-H) -gene could be used to detect C. trachomatis. The detection limit of each real-time PCR was 50 genome copies per reaction (Schaeffer, 2008). However, difficulties in distinguishing some of the C. trachomatis sero-variants by this method were acknowledged by the authors. If implemented on a large scale, the costs of RT-PCR diagnosis for C. trachomatis and time to diagnosis may, unfortunately, become prohibitive for the underserved populations. Therefore, a rapid and affordable point of care analysis of patient-derived specimens can revolutionize the field of diagnosis of genital chlamydial infections. Our technology will provide physicians with an inexpensive simple assay and unambiguous readout that will suggest, or rule out, the necessity of further treatment right in the physician's office. In summary, the assay-device combination proposed here will reach the underserved populations who can be much more fully protected when enabled with point of care diagnosis of genital chlamydial infections.