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Jun 2, 2014

Easing the Transition to Automated NGS Sample Preparation

By attending to several key factors, laboratories can increase throughput, achieve data consistency, and maximize walkaway time.

  • Any laboratory that performs next-generation sequencing (NGS) must accept certain burdens such as the burdens of sample preparation. This includes DNA/RNA isolation, adapter ligation, multiple rounds of purification, quality control, and (potentially) target enrichment. These sample preparation steps consist of a series of complex, time-consuming, and error-prone chores.

    Because NGS sample preparation is so demanding, it can severely limit productivity, particularly in laboratories that continue to rely on manual techniques. Many laboratories, however, have the option of transitioning to automated NGS sample preparation. This option may appear increasingly attractive as the cost of sequencing falls and the throughput of the next-generation sequencers rises.

    By automating NGS sample preparation, laboratories can increase throughput and boost productivity. They can also reduce variability, enhancing their ability to take on applications that require consistent results. Finally, laboratories can streamline sample preparation, reducing errors and worker fatigue, and leaving researchers more time to perform valuable tasks.

    To manage the transition from manual to automated NGS sample preparation, a laboratory should be clear about its particular needs. For example, does it need a compact system, or can it accommodate something larger—or would it prefer a scalable solution? Does the laboratory present any less-than-optimal environmental conditions? Will it need to manage multiple processes or coordinate operations across multiple locations? And, finally, is it a candidate for a dedicated, single-purpose instrument or a system capable of adapting to different processes?

    All these questions are addressed in the sections below. Additional considerations—pertaining less to the implementation and more to the optimization of automated NGS sample preparation—are addressed in this article’s sequel.

  • Platform Size

    Click Image To Enlarge +
    Targeted resequencing was carried out before and after automation by means of an Agilent Bravo Automated Liquid Handling Platform, which dispensed SureSelectXT reagents. Note that automation increased throughput nearly 10-fold while reducing hands-on time about 90%.

    A compact system is ideal for laboratories with bench space restrictions and/or tight budgets. It provides the benefit of reduced “hands on” time by automating the most tedious liquid handling steps (Table 1). A compact system, however, may need frequent “walk ups” so that operators may change tips boxes or introduce reagents. A larger system provides more walkaway time but may have additional limitations due to height. A 96-channel pipetting head on a compact system immediately enables a significant increase in throughput. Ideally, a laboratory should be able to start small if needed and then add solutions for increased walkaway time (Figure).

  • Click Image To Enlarge +
    Figure.

    Figure. Agilent platforms are designed to keep pace with changing laboratory needs. A platform that combines the Bravo Automated Liquid Handling Platform (left) can provide high throughput and improved reproducibility with a small footprint. Extending this platform by adding the BenchCel 4R Microplate Handler and the Labware MiniHub (right) can add the benefit of increased walkaway time. The PlateLoc Thermal Microplate Sealer (middle) provides automated sealing for improving PCR efficiency. In either configuration, NGS Bravo is equipped with accessories for heating, cooling, shaking, and magnetic separation.

  • Multiple Systems for Throughput and Consistency

    Laboratories may require ultra-high throughput, or they may need to carry out several different processes at the same time. (For example, laboratories have to prepare samples for different sequencers, or they may have to use different reagents for different applications.) Such laboratories may require multiple systems.

    Some laboratories may be doing pre- and post-PCR applications in separate labs. Typically, library preparation is done in the pre-PCR area and has more pipetting steps. To carry out this task, a larger system accommodating tip boxes and reagent plates may be more appropriate. In Post-PCR, however, a compact system might suffice, particularly when space or budget is constrained.

    For groups with multiple labs, having a similar set up in all labs helps drive data consistency. It also allows users to train each other and collaborate seamlessly, using the same kits and protocols.

  • Environment

    Ambient temperature should be maintained around 22–27°C. Very low temperature in labs can affect some steps in sample preparation, in which case slight warming of reagents may help. Care needs to be taken to prevent warming from increasing rates of evaporation. The automation set up should not be kept directly under air vents.

    If dust and other contaminants pose an issue, a containment system, such as a simple enclosure or one with HEPA filters, is recommended for the set up. Several containment systems are commercially available.

  • Flexibility

    Some automation systems are specifically designed to carry out one process or work with only one type of reagent. Such a system may work for a laboratory that does not expect to change its needs over the system’s lifetime. However for most laboratories, it is important that to have equipment that is able to keep pace with their changing needs. These laboratories should deploy equipment that is able to accommodate a variety of labware and accessories for temperature control, shaking, vacuum, magnetic bead separation, etc.

    Please see this article’s sequel for additional considerations affecting the success of automation. Also, be sure to check out the infographic "NGS Sample Prep—Manual vs. Automated".



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