Automated liquid handling for every lab

As automated liquid handling technology improves, could it move from specialist screening tool to routine lab staple? As he sums up the technology so far, Joby Jenkins finds some answers…

The ability to move liquids around the laboratory in carefully defined volumes is a key component much research, traditionally achieved using small, single- or multi-channel pipettes, operated by hand. However, liquid handling in this way can be impractical, monotonous and time-consuming in most situations, leading to inaccuracy and inconsistency as researchers get fatigued. These issues are compounded as the number of samples and pipetting steps increase, until eventually it becomes impossible to achieve reliable results in a reasonable time frame using manual pipetting. During the late eighties and early nineties, the increased throughput needs of screening laboratories for drug discovery exacerbated this problem, and drove the need for a robust, reliable solution. Automated liquid handling was scientists’ saving grace.

Although automated liquid handling was pioneered in those drug discovery laboratories, almost all life scientists are now under pressure to run larger assays that investigate more samples and biological targets in less time. This is creating an environment where many more researchers can benefit from automated liquid handling, which offers a cost-effective way to boost productivity and efficiency across a wide range of academic and industrial research applications.

An automated liquid handling solution should be quicker, more accurate and offer greater consistency than the same pipetting operations previously carried out by hand. Ideally, it should also be able to work with a range of volumes, applications and plate types and tolerate varying liquid viscosity and surface tension, while also successfully avoiding common liquid handling problems such as clogging, foaming, uneven dispensing and cross contamination (see Chai et al. 2013 for review1). To meet these needs, the last few decades have seen the development of ever-more powerful and flexible liquid handling instrumentation. This has required significant invention on multiple levels, including new instrument design, the introduction of software platforms capable of combining intuitive and flexible control with ease-of-use, and novel engineering solutions that increase pipetting accuracy at low volumes.

Current automated dispensing solutions employ a range of innovative engineering approaches to allow the fine control of liquid dispensing, primarily utilising either contact or non-contact methodologies. Both are designed to overcome the physical challenges imposed by surface adhesion, allowing even small volume droplets to be dispensed into the vessel of choice. As the name would suggest, contact-based dispensing requires that the liquid being dispensed makes contact with the target substrate, by physically moving the dispensing tip to the target. The dispensing channel is then withdrawn, with surface tension holding the liquid in place at the target location.

In contrast, non-contact dispensing uses additional force to eject the liquid from the dispensing tip or vessel and onto the target substrate, usually achieved via solenoid, piezoelectric or acoustic technologies (although other approaches do exist, see Kong et al 2012 for review2). Solenoid-based devices use positive-displacement controlled by a solenoid valve, which can be manipulated via an electronic current to make it open, close or redirect liquid flow. Piezoelectric dispensers are composed of capillary tubes made of quartz or steel, which are surrounded by a piezoelectric crystal collar. This collar contracts when a voltage is applied, thereby applying pressure to the capillary and forcing a small amount of liquid out of the dispensing tip. Lastly, acoustic dispensers use sound energy to propel small droplets onto the target substrate, without requiring direct contact with it.

Regardless of technological approach, viscosity is perhaps the biggest challenge faced by automatic liquid dispensing technologies. Poorly managed, it can lead to serious inaccuracies and a lack of consistency between droplets, especially when working at the low volumes required for modern assay optimisation, as solutions composed of water, oil or DMSO behave very differently at nL volumes. Therefore, labs requiring application flexibility would be well-advised to select a liquid handler capable of working reproducibly across a range of liquid types.

[caption id="attachment_34441" align="aligncenter" width="430" caption="utilising TTP Labtech’s mosquito automated liquid handler (in this case, in combination with the Gateway cloning system). A) BP clonase reaction, B) LR clonase reaction."][/caption]

Much of the early development of automated liquid handling was driven by the emergence of large and complex drug screening protocols designed to increase hit-to-lead success. Using such systems, it is now possible for high throughput screening laboratories to investigate the suitability of millions of drug candidates quickly, efficiently and accurately. The approach also brings with it many additional benefits, such as cost reductions through assay miniaturisation. By increasing the number of wells per plate (for example, via the use of high-density formats such as 384- and 1,536-well) and reducing the volumes used in each well, drug discovery researchers can screen more candidates in each assay, providing a greater amount of data per screen.

In the competitive world of drug development, the ability to cost-effectively screen large banks of compounds increases the chances of identifying high value drug candidates. As such, automated liquid handling is now common place for plate replication and serial dilutions in screening labs. However, the need for high throughput protocols now extends into many other application areas, including the preparation of stock solutions, serial dilution/dose response, plate-to-plate transfer, compound screening and hit-picking, biological assay setup and optimisation, nucleic acid extraction, PCR preparation, protein crystallisation and cell culture seeding/handling.

During the early days of automated liquid handling, the approach was considered unnecessary or cost-prohibitive by many research laboratories, particularly those not carrying out high throughput screening assays. However, the extensive accuracy, speed and cost benefits provided by automated systems, combined with an ever increasing burden on researchers of all disciplines to turn around a great number of assays in a cost-effective and time-efficient way, has led many to revaluate their position.

Much of this need has been driven by the emergence of the ‘omic’ era, with academic and industrial scientists now routinely carrying out large assays across entire genomes, proteomes and transcriptomes using approaches such as microarrays, qPCR and next generation sequencing (NGS). While such studies can provide a large amount of interesting and relevant data, they bring with them significant practical challenges, one of which is liquid handling. As such, more and more research teams are turning towards automated systems, as they allow scientists to focus more of their time and energy on analysing their data, rather than creating reaction mixtures, pipetting samples or performing many rounds of laborious assay optimisation by hand. In addition, automated liquid handling allows researchers to optimise sample use by carrying out more reactions, in less time and with greater consistency.

The increasing number and diversity of applications requiring access to automated liquid handling has stimulated the development of flexible systems capable of working across a wide range of dispensing volumes and assay types. This includes extended dispensing ranges across several orders of magnitude, as well as compatibility with multiple plate formats, such as those with 96, 384 or 1536 wells. Recent years have seen the development of instruments capable of meeting these requirements, allowing automated liquid handling to revolutionise workflows across a range of common molecular biology applications.

One area where automated liquid handling is providing significant benefits is for NGS analysis, by improving the cost-effectiveness and efficiency of sample preparation. Here, the use of automated instruments enables the reduction of reaction volumes, significantly lowering the cost-per-assay of using expensive reagents (for example, tagmentation enzymes). One recent proof-of-concept study carried out using a TTP Labtech liquid handler showed that overall assay volumes could be reduced from 50µL to 5µL or even 1µL, without comprising on accuracy and precision3. Such a reduction offers significant savings on the amount of genomic DNA used and reagent costs. Furthermore, the gentle pipetting action employed by this automated system reduced the potential for frothing (as can be the case with viscous enzyme mixes,) thus minimising the risk of unwanted DNA fragmentation, offering greater accuracy and potentially simpler data analysis.

Reaction miniaturisation using automated liquid handling is also providing significant cost savings for scientists carrying out DNA cloning using systems such as the Gateway cloning approach from Life Technologies. In the case of this application, viscous enzyme solutions can be pipetted accurately and reproducibly using low volume, positive displacement liquid handling solutions, enabling effective cloning efficiency to be maintained even when employing tenfold less enzyme per reaction (Figure 1).

In many respects, such assay optimisations are the tip of the iceberg, with low volume dispensing using automated systems likely to offer similar cost savings across a wide variety of other common molecular biology applications. These include ligation, DNA digestion, DNA/RNA purification, qPCR, cDNA synthesis and many more. However, many of these reactions require the accurate dispensing of liquids with a range of viscosities, so it is important to select an automated liquid handler flexible enough to perform reproducibly regardless of liquid viscosity or application.

Automated liquid handling instruments reduce processing time, decrease sample contamination, and increase accuracy across a range of assays, from compound screening through to NGS and protein crystallisation. These attributes release researchers from long, repetitive, laborious tasks, and free up time for analysis and thought. Many of the systems available on the market today are easy-to-use and provide automated pipetting for a range of assay types and sizes. Many are also easily integrated into existing instrumentation, enhancing laboratory throughput and efficiency.

As low volume pipetting accuracy using even viscous reagents continues to improve, the volume of reagents required on a per-reaction basis will drop even lower. This will further facilitate the technology’s rapid transition from a specialist screening tool to an essential part of many laboratory workflows, helping to drive discovery well into the future.

References: 1. Chai SC, Goktug AN, Cui J, Low J and Chen T (2013). Practical Considerations of Liquid Handling Devices in Drug Discovery. Book Chapter, “Drug Discovery”, edited by Hany A. El-Shemy, ISBN 978-953-51-0906-8 2. Kong F, Yuan L, Zheng YF and Chen W (2012). Automatic Liquid Handling for Life Science: A Critical Review of the Current State of the Art. Journal of Laboratory Automation 17(3): pp 169-85 3. Jenkins J (2013). A simple and automated solution for miniaturising reaction volumes for Nextera NGS sample preparation. TTP Labtech Application Note: http://discover.ttplabtech.com/DLMOSNGSappnoteNextera.html

Author: Joby Jenkins, Product Manager at TTP Labtech