Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, Wisconsin.
Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, Wisconsin.
J Mol Diagn. 2014 May;16(3):297-304. doi: 10.1016/j.jmoldx.2014.01.004. Epub 2014 Mar 6.
The monitoring of viral load is critical for proper management of antiretroviral therapy for HIV-positive patients. Unfortunately, in the developing world, significant economic and geographical barriers exist, limiting access to this test. The complexity of current viral load assays makes them expensive and their access limited to advanced facilities. We attempted to address these limitations by replacing conventional RNA extraction, one of the essential processes in viral load quantitation, with a simplified technique known as immiscible filtration assisted by surface tension (IFAST). Furthermore, these devices were produced via the embossing of wax, enabling local populations to produce and dispose of their own devices with minimal training or infrastructure, potentially reducing the total assay cost. In addition, IFAST can be used to reduce cold chain dependence during transportation. Viral RNA extracted from raw samples stored at 37°C for 1 week exhibited nearly complete degradation. However, IFAST-purified RNA could be stored at 37°C for 1 week without significant loss. These data suggest that RNA isolated at the point of care (eg, in a rural clinic) via IFAST could be shipped to a central laboratory for quantitative RT-PCR without a cold chain. Using this technology, we have demonstrated accurate and repeatable measurements of viral load on samples with as low as 50 copies per milliliter of sample.
病毒载量监测对于 HIV 阳性患者的抗逆转录病毒治疗的合理管理至关重要。不幸的是，在发展中国家，存在着巨大的经济和地理障碍，限制了人们获得这种检测的机会。目前病毒载量检测方法的复杂性使得它们昂贵，而且只有在先进的设施中才能获得。我们试图通过用一种称为表面张力辅助不混溶过滤（IFAST）的简化技术来取代常规的 RNA 提取（病毒载量定量的一个必要过程）来解决这些限制。此外，这些设备是通过蜡压印制成的，使当地居民能够在最少的培训或基础设施的情况下生产和处理自己的设备，从而有可能降低总检测成本。此外，IFAST 还可以用于减少运输过程中对冷链的依赖。在 37°C 下储存 1 周的原始样本中提取的病毒 RNA 几乎完全降解。然而，IFAST 纯化的 RNA 可以在 37°C 下储存 1 周而不会有明显的损失。这些数据表明，通过 IFAST 在护理点（例如，在农村诊所）分离的 RNA 可以在没有冷链的情况下运送到中央实验室进行定量 RT-PCR。使用这项技术，我们已经证明了在低至每毫升样本 50 个拷贝的样本中，都可以进行准确且可重复的病毒载量测量。
