Automation has always been driven by the need to increase accuracy and speed, and minimize human intervention. This need has escalated as a result of the COVID-19 pandemic with laboratories striving hard to develop diagnostics and therapeutics to fight the virus, while limiting the number of people doing the work. Finding ways to automate nucleic acid purification to increase throughput, reproducibility, and accuracy, while also processing samples with a high degree of purity, has been crucial in this effort.
According to Luke Roenneburg, senior strategic portfolio manager, sample prep at Promega, new systems minimize human touch points by incorporating higher levels of automation for the entire workflow and relying on software to reduce human error. Promega is developing modular instruments that work together through connected software to address complete workflows from sample preparation to extraction to downstream preparation of purified nucleic acids for various applications.
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Markus Sprenger-Haussels, VP, head of sample technologies product development, life sciences at QIAGEN, adds that nucleic acid purification is evolving in different directions. On one hand, are the more specialized procedures tailor-made to achieve the best possible performance with difficult sample types, such as feces, soil, or plant material, and less focused on speed or ease-of-use. “Here the main challenge is to get rid of inhibitors and interfering substances, which could have an impact on potentially expensive downstream assays,” he says. On the other hand, are closed workflows in which a nucleic acid extraction module is fully integrated into microfluidic, sample-to-insight workflows, consisting of all relevant steps from sample disruption, nucleic acid extraction, amplification, and/or detection. “In these workflows the nucleic acid isolation can fully be tweaked toward one dedicated sample type and downstream reaction, and the driving force is speed and ease-of-use,” says Sprenger-Haussels.
“In this race for the fastest and most convenient workflow, we just saw the launch of a one-step liquid nucleic acid extraction for high-throughput SARS-CoV-2 testing (e.g. QIAprep&),” says Sprenger-Haussels. “Here, the extraction and amplification have been adjusted to work perfectly together. Each part alone does not function any better, but if combined, the performance can compete with any full-blown extraction and amplification protocol using only a fraction of the time.”
Need for simplicity
The first thing to look at when automating nucleic acid purification is simplicity of use, explains Daniel Weissbart, product manager and technical support director, MP Biomedicals. “The installation of consumables, samples, and the launch of the program must be done quickly and without hesitation.”
Key Questions to Ask
- Does it work for your sample?
- What is the cost of analysis per sample?
- How good is the quality, yield, and reproducibility of extraction?
- How simple is it to use?
- How quickly can it integrate into your workflow?
- Does it minimize sample handling and preparation?
- Can it handle sample versatility and scalability?
- Does it prevent cross-contamination?
- Does it offer flexibility in the number and volume of samples to be processed?
- Is it scalable and customizable to meet your needs?
- Does it offer ready-to-run sample purification kits?
- Is it eco-friendly and minimize waste production?
- Is it difficult to maintain?
- Can you run some samples before purchase?
- What kind of warranty and after-sales service does the vendor provide?
While automation provides tremendous time and cost savings, it can also add complexity depending on how well it fits into the workflow. Humans can adapt protocols and workflows to what’s needed but it becomes difficult to make changes once processes are automated. “Many automated platforms are closed systems and restrict the end-user to a specific set of reagents and kits,” says Celia Landers, product segment head of molecular prep and PCR at MilliporeSigma. “Hence, investments in any instrumentation in the lab, especially automation of nucleic acid purification, should be considered with long-term research plans in mind to maximize flexibility, including with sample types, potential analytes of interest, and changes in throughput.”
Need for speed
The SARS-CoV-2 pandemic forced laboratories to get, store, run, and analyze samples in a matter of days. It honed in on instruments and processes that were easy to use and adopt, with minimal knowledge and expertise. “We focus on easy-to-use, benchtop devices for nucleic acid extraction paired with application-specific reagents that can be incorporated into laboratory workflows rapidly, which technicians can use to quickly process samples with minimal training,” says Roenneburg.
“We have seen downstream applications becoming less sensitive to contaminants and inhibitors and also an increased ability to analyze targets that allow extractions to focus on reducing overall time to obtain purified nucleic acids.” There is also an increasing need to obtain more data from a single sample with multiplex assays and multi-modal sequencing panels. “A key driver in any new extraction chemistry that Promega develops is going to look for ways in which the number of manual sample preparation steps can be reduced or removed altogether with direct sample extraction,” adds Roenneburg.
MilliporeSigma’s new DNA and RNA purification kits use a negative chromatography method based on size exclusion for the purification of genomic DNA (gDNA) from mammalian cells, blood, tissue, and plant tissue samples, as well as for DNA and RNA cleanup procedures. “While traditional silica-based spin prep kits require multiple, tedious wash and spin steps with lots of hands-on manipulation and little downtime, these new purification columns efficiently absorb and retain sample contaminants while allowing nucleic acids to flow through the column, greatly reducing the number of steps and materials required for purification,” says Landers. The pandemic-related shortage in silica has also prompted the use of new eco-friendly alternatives to silica.
Need for quality
Isolation of high-quality nucleic acids is critical for sensitive downstream applications such as in the life sciences, diagnostics, sequencing, forensics, and genetic engineering. Hence, choosing the right nucleic acid isolation and purification kits for automated nucleic acid extraction systems is very important. Véronique Karsten, product manager, molecular biology at MP Biomedicals, reports that their MPure-12 platform incorporates a uniquely designed polygon reaction chamber ensuring better extraction performance with no risk of carry-over and cross-contamination, as well as no tip clogs and partial liquid transfers. “The barcode reader also allows for quick set-up of protocols by scanning the pre-programmed protocol cards and a UV lamp facilitates decontamination between runs, which guarantees purity of samples.”
Image: The MPure-12™ Automated Nucleic Acid Purification System is a fully automated and integrated platform to purify nucleic acids for downstream applications. Image courtesy of MP Biomedicals
Need for flexibility
According to Landers, customers are increasingly interested in difficult-to-work-with sample types, such as formalin-fixed paraffin-embedded (FFPE) samples or fatty, fibrous tissues, and precious samples like biopsies. “I think there will be a continuing evolution of kits with specialized lysis and inhibitor removal steps for difficult samples for sensitive downstream applications like digital PCR or next-generation sequencing. We will also continue to see more multi-analyte purification kits as customers continue to do more with less sample,” she says.
There is a growing need for automating nucleic acid purification to recover DNA and RNA from the toughest samples used in microbiome work such as soil and stool samples, says Karsten. “The Mpure-12 benchtop automated system uses magnetic bead separation technology and is supplied with a wide range of sample-specific pre-packaged reagent kits. It has been used for the isolation of high-quality viral RNA for accurate and sensitive SARS-CoV-2 detection in swabs via quantitative reverse transcriptase PCR (qRT-PCR).” The emergence of new sequencing technologies has created new demands in nucleic acid extraction. “Third-generation sequencers and whole genome sequencing have created a need for extraction technologies that provide more intact and larger nucleic acid fragments,” adds Roenneburg.
Need for innovation
Innovation and customization are driving the changes in nucleic acid isolation and purification workflows. “We are looking into multi-modality applications, analyzing DNA and RNA from the same sample, DNA sequencing with increased reading length requiring high molecular weight DNA extraction, applications like microbiome analysis demanding bias-free extraction efficiency (extract all bacteria—gram positive or negative—with the same efficiency to fully reflect the population), reducing environmental DNA contaminations for low biomass applications, and the environmental footprint of nucleic acid extraction (e.g. reduction of plastic waste), just to mention a few activity fields,” says Sprenger-Haussels. Diagnostic and forensic laboratories as well as academic research centers are calling for the development of a compact, portable, field-deployable, nucleic extraction system that will be valuable for both environmental microbiology, as well as health care diagnostics, says Karsten.
Innovation is also driving changes in reagents and columns materials. “Nanometer-sized paramagnetic particles, with different coating shells including monolayer ligands, polymers, combinations of polymers, biomolecules such as phospholipids and carbohydrates, and inorganic materials like silica, provide not only the stability to nanoparticles in solution but also help in binding several biological ligands on the nanoparticle surface, as needed for medical applications,” notes Weissbart.
“The field of nucleic acid applications is so huge and diverse, there is simply no one-size-fits-all solution,” says Sprenger-Haussels. “Depending on the requirement of each application we can design the best possible workflow, spanning the entire range: any sample type, any target nucleic acid, any throughput—manual or automated and combining any bioinformatic pipeline—to achieve the best outcome.”
What to Look For
- Less manual intervention
- High-throughput and speed
- Simplified workflow
- Multiplex operation
- Low-cost extractions
- Less sample preparation
- High purity, minimal cross-contamination
- Pre-programmed protocols and pre-set reagents
- Ability to customize protocols
- Compact, portable systems for field applications