HID NIMBUS Presto System for Automated Sample Purification

The deck-mounted KingFisher Presto gently and efficiently isolates and purifies nucleic acids from a range of sample types for use in sensitive downstream analyses such as DNA quantification and STR amplification. The process utilizes magnetic particle-based technology to facilitate high-quality nucleic acid purification. When coupled with PrepFiler or PrepFiler BTA Automated extraction chemistries, DNA binds to the magnetic particles, cellular debris/inhibitors are removed, and the bound DNA is eluted through the movement of the magnetic particles across reagent plates (Figure 1).

A dual-position turntable allows processing of one plate while the integrated automated liquid handler simultaneously prepares and transports a second plate, without the need for manual intervention. The automated liquid handler's patented pipette tip technology creates a tight seal for superior liquid measurement accuracy and precision without tip distortion or aerosol generation during tip pick-up and ejection, so plates are prepared accurately and efficiently. The flexible format includes integrated tools like labware grippers to automatically transport plates and around the deck and a barcode scanner to track samples.

The HID NIMBUS Presto System combined with the PrepFiler Automated chemistries is able to repeatedly and consistently isolate or extract actionable amounts of DNA from limited and challenging forensic sample types. Dilution series of blood and saliva spotted on swabs were processed with the PrepFiler 300 μL and 500 μL lysis protocols on the HID NIMBUS Presto, and on the AutoMate Express with the standard PrepFiler protocol to assess comparability. (Figure 2). Similar results were seen with the PrepFiler BTA chemistry (data not shown). Internal Positive Control (IPC) CTs from the quantification assay indicated that all samples were free of inhibitory factors (data not shown).

The HID NIMBUS Presto System delivers DNA from challenging samples and is capable of generating full genetic profiles for use in forensic casework analysis. A variety of known and non-probative casework-type samples were used to evaluate the ability of the HID NIMBUS Presto System to extract inhibitor-free DNA of the highest quantity and quality possible from common human identification sample types; when noted, results from same samples extracted with AutoMate Express for comparative purposes are shown (Figure 3).

Figure 3. 3500xL Genetic Analyzer electropherograms of mock case-type samples extracted on HID NIMBUS Presto unless otherwise noted and amplified with the GlobalFiler IQC PCR Amplification Kit showing an overlay of all dye channels. The Y-axis is adjusted individually per sample as indicated. From upper panel to lower: Swab of drink can rim extracted with PrepFiler 300 μL lysis protocol; swab of dried blood (2.5 μL) on surface extracted with PrepFiler 500 μL lysis protocol; 1 μL of dried saliva on tape extracted with PrepFiler BTA; 50 mg of slightly degraded bone sample extracted with PrepFiler BTA; 50 mg of slightly degraded bone sample extracted with PrepFiler BTA processed on AutoMate Express.

The automated liquid handling robot includes a patented pipette tip technology that creates a tight seal for superior liquid measurement accuracy and precision without tip distortion and aerosol generation during tip pick-up and ejection, thus limiting cross contamination.

The checkerboard pattern in Figure 4, alternating high-level control single-donor DNA input samples with extraction reagent blanks, confirms that the HID NIMBUS Presto System does not introduce cross-contamination during automated extraction (PrepFiler BTA and PrepFiler 500 μL lysis protocols were tested).  

Figure 4. (Top panel) Sample input orientation in processing plate for each contamination study. Reagent blanks are indicated with pink shading and were placed in a checkerboard pattern around high-level control DNA (500 ng in PrepFiler 500 μL lysis protocol study 300 ng in PrepFiler BTA study,) input samples. (Middle panel) Electropherogram overlay of all 36 reagent blanks, displaying all dye channels, purified on the HID NIMBUS Presto System with PrepFiler 500 μL lysis protocol, then amplified with the GlobalFiler IQC PCR Amplification Kit (15 μL of eluate, 29 cycles). The Y-axis is scaled to 200 RFU, One Out-of-Marker-Range (OMR) doublet, one CE artifact spike and one in-bin peak (51 RFU) at an allele not shared with Donor DNA genotype is present above 50 RFU. Blue-channel peaks at far ends of either side of the EPG are the Internal Quality Control (IQC) peaks which are intrinsic to the kit. (Lower panel) Electropherogram overlay of all 36 reagent blanks, displaying all dye channels, purified on the HID NIMBUS® Presto System with PrepFiler BTA protocol, then amplified with the GlobalFiler IQC PCR Amplification Kit (15 μL of eluate, 29 cycles). The Y-axis is scaled to 200 RFU, One CE artifact spike and one in-bin peak (64 RFU) at an allele not shared with Donor DNA genotype is present above 50 RFU. Blue-channel peaks at far ends of either side of the EPG are the Internal Quality Control (IQC) peaks which are intrinsic to the kit. Absence of cross-contamination during extraction process is evidenced by lack of peaks matching donor DNA genotype in any of the reagent blanks.

Since 2007, the Human Identification Professional Services (HPS) team has been helping customers successfully navigate the validation process required to bring new technologies into operation. HPS has completed over 700 validation and integration projects in more than 39 countries worldwide. With a dedicated team of validation specialists, each with forensic experience, HPS provides forensic laboratories with comprehensive validations and implementation support, both vital for bringing new technologies into operation. Using a consultative approach, the HPS team designs a service that meets most technical and budgetary needs. HPS has designed service offerings specifically for the HID NIMBUS Presto System with the PrepFiler Automated Forensic DNA Extraction Kit.

Thermo Fisher Scientific performed developmental validation experiments in accordance with the DNA Advisory Board (DAB) Quality Assurance Standards (July 1, 2020) and guidelines from the Scientific Working Group on DNA Analysis Methods (SWGDAM, December 5, 2016) to evaluate the performance of the HID NIMBUS Presto System with PrepFiler and PrepFiler BTA Automated Forensic DNA Extraction Kits.

Thermo Fisher Scientific has validated the instrument -reagent system for use in Human Identification testing.