Department of Chemistry, University of Virginia, Charlottesville, VA, USA.
New England Biolabs, Ipswich, MA, USA.
J Forensic Sci. 2021 May;66(3):1033-1041. doi: 10.1111/1556-4029.14682. Epub 2021 Feb 9.
Accurate presumptive and confirmatory test use for forensic body fluid identification is essential for gaining contextual information for crime scene investigators. Loop-mediated isothermal amplification (LAMP) is an ideal method for forensic body fluid identification because it is highly specific and generates multi-sized amplicon DNA, and successful amplification results can be read out colorimetrically. Here, we show preliminary data on a LAMP method that rapidly identifies body fluids including venous blood, semen, and saliva, based on colorimetric response and image analysis. The method is designed for easy implementation into forensic casework protocols with minimal disruption to DNA analysis. LAMP naturally increases target specificity due to the use of multiple primers for one target and mRNA targets were used for tissue and human specificity. With colorimetric detection as an inherent part of LAMP, samples that are positive or negative for any of the body fluids are readily identified by image capture and analysis, thus eliminating subjectivity. Results show by using the 3D-printed imaging system specific color ranges can be set for easy determination of body fluids. The resulting color change can be seen in <30 min using a universal temperature and primer concentration for all body fluids. This simple method and imaging system allow for minimal hands-on time with objective image analysis and presents a pathway for creating a new potential method for forensic body fluid identification.
准确使用法医体液鉴定的初步和确认测试对于犯罪现场调查人员获取背景信息至关重要。环介导等温扩增 (LAMP) 是法医体液鉴定的理想方法，因为它具有高度特异性并产生多大小的扩增子 DNA，并且可以通过比色法读取成功的扩增结果。在这里，我们展示了一种基于比色响应和图像分析的快速鉴定包括静脉血、精液和唾液在内的体液的 LAMP 方法的初步数据。该方法旨在轻松地融入法医工作协议中，对 DNA 分析的干扰最小。由于 LAMP 对一个靶标使用多个引物，因此自然会增加靶标特异性，并且使用 mRNA 靶标来提高组织和人体特异性。由于比色检测是 LAMP 的固有部分，因此可以通过图像捕获和分析轻松识别任何体液呈阳性或阴性的样本，从而消除主观性。结果表明，使用 3D 打印成像系统可以为容易确定体液设置特定的颜色范围。使用所有体液的通用温度和引物浓度，在 <30 分钟内即可看到这种颜色变化。这种简单的方法和成像系统允许进行最少的手动操作，并进行客观的图像分析，为创建法医体液鉴定的新潜在方法提供了途径。
