Sep 29 2021
Extracting and Purifying DNA from Challenging Calcified and Adhesive-Based Samples in Parallel Using a Walkaway Robotic System
Calcified samples or those that include an adhesive substrate may require more robust processing compared to routine forensic sample types. When performed manually, DNA extraction and purification of these challenging samples can impede overall laboratory efficiency and invite risks of human error and variability. A novel combination of assay chemistry and robotic workstation aid robust, repeatable, and hands-free DNA extraction and purification of even the most challenging sample to ensure high yields of pure DNA for downstream analyses such as capillary electrophoresis or massively parallel sequencing. At the same time, it facilitates highthroughput and parallel processing, up to 96 batched samples in volumes ranging from 50 – 5000 µL, to enhance productivity in the lab.
Written by: Kevin Miller, Hamilton Company
Being a forensic scientist myself, I am always interested in solving the pain points encountered within forensic workflows. In this instance, the ID NIMBUS Presto offers the ability to rapidly isolate DNA from up to 96 evidentiary samples at a time, using an automated system with a relatively small footprint. Hamilton Company and Thermo Fisher Scientific have partnered to enable forensic scientists to obtain DNA from even the most challenging evidentiary samples by providing reliable, hands-free extraction solutions. Project collaborators included Matthew Ludeman, Angela Chen, Ellen Crone, and Julio Mulero, all from Thermo Fisher Scientific, and William Hutchens from Hamilton Company.
We developed a novel automated liquid handling workstation containing an integrated KingFisher™ Presto unit, and evaluated its ability to purify DNA from commonly encountered forensic-type samples such as touch/trace, bone, tooth, and adhesive-containing materials. The ID NIMBUS® Presto assay-ready workstation is different from other automated forensic DNA extraction solutions, because magnetic rods inserted directly into the sample move DNA-containing magnetic beads from one clean reagent to the next rather than having reagents aspirated and dispensed around a cluster of beads held to the bottom or side of a single sample vessel. These actions, together with the ability of the robot to independently prepare sample plates with reagents ahead of when they are needed, make the ID NIMBUS Presto incredibly efficient. In fact, the workflow is incredibly simple (Lyse > Bind > Wash > Elute) and DNA is able to be purified from up to 96 evidentiary samples in as little as 90 minutes.
We used the PrepFiler™ BTA Automated Forensic DNA Extraction Kit for purifying DNA from common human identification sample types using the ID NIMBUS Presto. Studies included sensitivity, reproducibility, cross-contamination, and accuracy.
Our studies demonstrated that the ID NIMBUS Presto assay-ready workstation, using the PrepFiler BTA DNA Extraction Kit, delivers purified genomic human DNA from forensic-type samples that is highly suitable for downstream applications such as real-time quantitative PCR and PCR for STR profiling. The automated system:
- Provides highly reproducible and repeatable results over a wide range of inputs and with different sample input types, even when the sample input is a challenging one, such as bone.
- Is robust with forensic case-type samples and mock touch/trace samples.
- Delivers eluates that are consistently free of PCR inhibitors, as determined by the Quantifiler™ Trio DNA Quantification Kit IPC CT values and IQC peaks generated with the GlobalFiler™ IQC PCR Amplification Kit.
- Exhibits no detectable cross-contamination of human DNA at 175 RFU PAT, and only a low-level, infrequent apparent cross-contamination event at 50 RFU PAT.
- Is comparable to the lower throughput AutoMate Express™ Forensic DNA Extraction System when using the same BTA chemistry.
Going forward, I will continue to work with laboratories to automate their sexual assault evidence sample processing using the AutoLys STAR SAE, and with crime scene investigators who can use the AutoLys-A tube as an evidence collection device. When used together, the time from collection to answer is greatly reduced.
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