by Tine Thorbjoernsen and Helge Lubenow

Sample collection and sample processing — preanalytical procedures — are the decisive steps of the analytical process chain. No level of sophistication in the downstream detection can compensate for compromise at the start. Automation can help forensic investigators to efficiently process samples and enables high sample throughput and rapid data generation. Throughput, however, is not the only, and probably not even the most important factor for the increasing number of forensic laboratories that have started to implement automation. Robotic processing minimizes human interaction with samples and thus helps to control contamination by environmental DNA. Automation can be an important means by which to ensure that standard operating procedures are employed. Standardization helps to secure technical quality and reproducibility, and allows full compatibility of data generated, for example, by different investigators at different locations or at different times. Data management interfacing with robotics makes complete documentation of the process and workflow easier and helps implement a chain of custody.

Automated purification of DNA from a variety of forensic samples

The EZ1 DNA Investigator Kit and Card provide: efficient yields — even from trace casework samples; high signal-to-noise ratios; easy handling of swabs, blood discs and other solid samples, which can be processed directly on the EZ1 workstation; a range of optimized protocols, including larger starting volumes (500 μl) for more dilute samples and normalization for uniform yields; and optional use of carrier RNA for increased extraction efficiency with sample types with low DNA content.

Automated purification of genomic DNA on the EZ1 workstation is based on silica-coated magnetic particles. Magnetic-particle technology eliminates tedious centrifugation steps, and allows for rapid automated processing. The workstation purifies 1–6 samples in a single run in just over 20 minutes. The workstation door and use of sealed reagent cartridges reduce the chances of contamination from the environment or operator.

In accordance with the manufacturer’s ISO-certified Quality Management System, each lot of EZ1 kits is tested against predetermined specifications to ensure consistent product quality. Every kit is supplied with a detailed certificate of analysis documenting the quality control tests and kit performance. All kit reagents are manufactured in a controlled environment and are monitored for the presence of contaminating human DNA.

The kit provides optimized chemistries for automated DNA purification. STR analysis shows high peaks with good signal-to-noise ratios and even peak distribution for sensitive detection (Figure 2).

In addition to the standard “trace samples” protocol application, a recently available “tip dance” protocol allows for fully automated DNA extraction from crime-scene samples without the need to remove the sample from the lysis reaction prior to the extraction procedure. This enables processing of solid materials, such as swabs, fabrics, blood discs, or cigarette butts, directly in the sample tube. There is no need for prior centrifugation to remove solid materials that could clog the tip. The “tip dance” protocol simplifies handling and, in some cases, increases DNA yield with more efficient extraction of the sample (Figure 3). In the “tip-dance” process, the pipetting tip of the robotic system moves back-and-forth relative to the worktable platform while pipetting and thus avoids clogging by the sample.

Large sample volume protocol

Normalization, set-up and handling

The pre-programmed card makes setting up the purification protocols as easy as making a credit card transaction. Pre-filled, sealed reagent cartridges in the EZ1 kit help to prevent human error during setup of the workstation and prevent contamination of buffers. By eliminating human error in reagent preparation and workstation setup, and by providing cross-contamination–free liquid handling, the workstation provides high levels of reliability and standardization.

About the authors

The EZ1 kit is intended for general laboratory use. No claim or representation is intended to provide information for the diagnosis, prevention, or treatment of a disease. The EZ1 card and the workstation are intended for research use. No claim or representation is intended to provide information for the diagnosis, prevention, or treatment of a disease.

More information is available from:

Figure 1. The BioRobot EZ1 workstation.

Figure 2. Improved performance in STR analysis. AmpFlSTR (Applera Corp. Norwalk, CT) control DNA (1 ng) was diluted in 200 μl buffer G2 and purified on the BioRobot EZ1 workstation. DNA was eluted in 50 μl water, and 10 μl (corresponding to 200 pg DNA) was used for STR analysis. A) DNA was purified using the EZ1 DNA Investigator Kit and the trace protocol on the EZ1 DNA Investigator Card. B) DNA was purified using the EZ1 DNA Tissue Kit and the trace protocol on the EZ1 DNA Forensic Card (Data kindly provided by B. Bayer and K. Anslinger, Institute of Legal Medicine, Ludwig Maximilian University, Munich, Germany).

Figure 3. Easy and efficient processing of solid materials with the “tip dance” protocol. Papers from cigarette butts or 3 paper discs per sample were spotted with 10 ng DNA per sample. After proteinase K digestion, half of the samples were processed using the EZ1 DNA Investigator Kit with the “tip dance” protocol, without removing solid materials from the sample tubes (Tip dance). Solid materials were removed from the remaining half of the samples, which were then processed using the EZ1 DNA Investigator Kit with the standard trace protocol (Trace). DNA yields were quantified by real-time, quantitative PCR.

Figure 4. Higher yields of concentrated DNA with the large-volume protocol. DNA was diluted in Buffer G2 to a final concentration of 50 pg/μl, and the indicated volumes were processed using EZ1 kit with the standard trace protocol (Trace) or the large-volume protocol (LV). All samples were eluted in 50 μl water, and 5 μl was quantified using real-time, quantitative PCR.