Optical tweezers as an effective tool for spermatozoa isolation from mixed forensic samples

Abstract

A single focus optical tweezer is formed when a laser beam is launched through a high numerical aperture immersion objective. This objective focuses the beam down to a diffraction-limited spot, which creates an optical trap where cells suspended in aqueous solutions can be held fixed. Spermatozoa, an often probative cell type in forensic investigations, can be captured inside this optical trap and dragged one by one across millimeter-length distances in order to create a cluster of cells which can be subsequently drawn up into a capillary for collection. Sperm cells are then ejected onto a sterile cover slip, counted, and transferred to a tube for DNA analysis workflow. The objective of this research was to optimize sperm cell collection for maximum DNA yield, and to determine the number of trapped sperm cells necessary to produce a full STR profile. A varying number of sperm cells from both a single-source semen sample and a mock sexual assault sample were isolated utilizing optical tweezers and processed using conventional STR analysis methods. Results demonstrated that approximately 50 trapped spermatozoa were required to obtain a consistently full DNA profile. A complete, single-source DNA profile was also achieved by isolating sperm cells via optical trapping from a mixture of sperm and vaginal epithelial cells. Based on these results, optical tweezers are a viable option for forensic applications such as separation of mixed populations of cells in forensic evidence.

Recent trends and developments in forensic DNA extraction

“…Optical trapping has previously been used to study sperm motility under various conditions (Nascimento, Botvinick, Shi, Durrant, & Berns, 2006; Tadir et al, 1989) and also applied to cell sorting on a microfluidic chip (Wang et al, 2011). This application was recently extended to selective isolation of sperm (Auka et al, 2019). The optical tweezer is a high‐density focused laser beam that uses an immersion objective lens on an inverted microscope to form an optical trap, capable of holding dielectric particles in the center (Ashkin, Dziedzic, Bjorkholm, & Chu, 1986).…”

“…The optical tweezer is a high‐density focused laser beam that uses an immersion objective lens on an inverted microscope to form an optical trap, capable of holding dielectric particles in the center (Ashkin, Dziedzic, Bjorkholm, & Chu, 1986). For the separation of sperm, a sperm is held in an optical trap, prior to transfer into a capillary for collection (Auka et al, 2019). Similar to LMD, 50 sperm are required to generate a high‐quality DNA profile.…”

Pushing the Boundaries: Forensic DNA Phenotyping Challenged by Single-Cell Sequencing

“…Mixture deconvolution is one of the most engaging topics in forensics. Over previous years different methods have been developed to assist the process, both prior to [ 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 ] and after, DNA typing of a mixed trace [ 36 , 37 , 38 ]. The DEPArray System, which uses a dielectrophoresis grid to isolate single cells, is a promising cell separation technology for forensic analysis.…”

“…The main motivation behind introducing this technology into forensics is the potential to improve mixture deconvolution workflows and interpretation methods. If a trace is a mixture of different biological materials (for example, sperm and epithelium), there are methods to separate cell pools to investigate them separately [ 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 ]. Problems arise when the mixed trace is too complex or if it consists of the same cell type, such as blood-blood mixtures [ 11 ].…”

DNA Transfer in Forensic Science: Recent Progress towards Meeting Challenges

“…Three studies continue to look at novel means of separating sperm cells from other cells to assist the interpretation of DNA profiles generated downstream. These include the application of optical tweezers [107], acoustic differential extraction technologies [108], and capillary zone electrophoresis [109]. Furthermore, using DEPArray™ technology (Menarini Silicon Biosystems, Bologna, Italy), Anslinger et al [110] demonstrated the ability to obtain complete DNA profiles from isolated single white cells from each of the two or three known contributors to mixed stains.…”