Morehouse Zachary P, Proctor Caleb M, Ryan Gabriella L, Nash Rodney J
Michigan State University College of Osteopathic Medicine, East Lansing, MI, USA.
Omni International Inc, A PerkinElmer Company, Kennesaw, GA, USA.
Bio Protoc. 2022 Mar 5;12(5):e4349. doi: 10.21769/BioProtoc.4349.
The impact of viral diseases on human health is becoming increasingly prevalent globally with the burden of disease being shared between resource-rich and poor areas. As seen in the global pandemic caused by SARS-CoV-2, there is a need to establish viral detection techniques applicable to resource-limited areas that provide sensitive and specific testing with a logistically conscious mindset. Herein, we describe a direct-to-PCR technology utilizing mechanical homogenization prior to viral PCR detection, which allows the user to bypass traditional RNA extraction techniques for accurate detection of human coronavirus. This methodology was validated , utilizing human coronavirus 229E (HCoV-229E), and then clinically, utilizing patient samples to test for SARS-CoV-2 infection. In this manuscript, we describe in detail the protocol utilized to determine the limit of detection for this methodology with testing of HCoV-229E.
病毒性疾病对人类健康的影响在全球范围内日益普遍，疾病负担在资源丰富和贫困地区之间分担。正如在由严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）引起的全球大流行中所看到的那样，需要建立适用于资源有限地区的病毒检测技术，这些技术要以具有后勤意识的思维方式提供灵敏且特异的检测。在此，我们描述了一种在病毒PCR检测之前利用机械匀浆的直接PCR技术，该技术允许用户绕过传统的RNA提取技术来准确检测人冠状病毒。这种方法利用人冠状病毒229E（HCoV-229E）进行了验证，然后在临床上利用患者样本检测SARS-CoV-2感染。在本手稿中，我们详细描述了通过检测HCoV-229E来确定该方法检测限所使用的方案。
