A Hybrid Approach to Liquid Handling


In order to develop an efficient and reproducible approach to liquid handling, researchers must be aware of the great diversity of liquid handling solutions available to them. The following article outlines existing solutions for manual and automated liquid handling, and proposes a hybrid approach that leverages the strengths of various technologies in the marketplace.



Manual Liquid Handling

Due to the number of liquid transfers required to perform molecular biology experiments, pipettes exist as one of the most ubiquitous symbols of daily laboratory work. However, due to the number of vendors in the market place and the wide range of products offered by each vendor, researchers often struggle to select the best pipette for their application. When selecting a pipette for a given transfer, important considerations include the physical properties of the liquid to be transferred, the volume of liquid to be transferred, the number of transfers required in a given workflow, and the accuracy and precision demanded in order to generate statistically meaningful and reproducible data.


Accuracy and Precision

When working with aqueous liquids, single channel, and fixed volume manual pipettes provide the highest degree of accuracy and precision. However, most researchers sacrifice accuracy and precision for ease-of-use, efficiency and speed. For most applications variable volume pipettes provide sufficient specifications to generate trustworthy results.


When selecting a variable volume pipette, the pipette utilizing the smallest piston, or capable of the smallest volume transfer, will yield the greatest accuracy due to the impact of the piston diameter on the area of the air cushion inside the shaft of the pipette. For instance, in the specifications below, the pipette with a volume range from 0.5 µL - 10 µL produces the most accurate and reproducible 10 µL dispense.

Pipette Model Accuracy at 10 µL Precision at 10 µL
0.5 µL – 10 µL 1.0% 0.4%
2 µL – 20 µL 1.5% 0.5%
10 µL – 100 µL 3.5% 1.0%


When performing higher throughput workflows, 8-, 12-, and even 96- channel pipettes provide increased efficiency. However, depending on the mechanics behind the parallel piston-operated channels, accuracy and precision will vary across products and vendors.


User-to-User Variability

It is well known that different users can produce varying pipetting results of the same experiment with the differences in the speed and force with which they press on the pipette plunger. To help remedy this, electronic air displacement pipettes have a motorized piston that consistently uses the same speed and force while dispensing.  This increases the reproducibility of liquid transfers, however other factors including the angle of the pipette, the depth of the tip underneath the surface of the liquid, the quality of pipette tips, the calibration of the pipette, and the physical properties of the liquid also largely impact liquid handling performance.


Air Displacement vs. Positive Displacement Pipetting

For aqueous samples, traditional air displacement pipetting provides sufficient accuracy and precision across all volumes. For problem liquids that have high densities, viscosities or vapor pressures, positive displacement pipetting ensures better pipetting performance through an approach by which a disposable piston directly contacts the liquid removing the need for an air cushion. This technology also provides benefits in the context of contamination, radioactivity, and corrosion.

Although the technologies employed by air displacement pipettes fail to provide reproducible liquid transfers in the case of certain liquids, a wider range of mutichannel solutions from 8- to 384- channel provide efficient dispensing in higher throughput workflows.



Automated Liquid Handling

Automated liquid handling benefits researchers by providing a means to remove all user-to-user variability from experimental procedures and by freeing up their time for more intellectual contributions in the lab. Since capabilities of automated liquid handling products vary greatly from simple plate stamps through full workflow automation, the following considerations provide some insights into the range of products available and provides a starting point to those considering automating their entire liquid handling workflow for a particular application.



The number of samples that researchers wish to process in a single run dictates the size of the liquid handling workstation required. At the low end of the throughput spectrum, personalized liquid handlers designed to process one plate at a time and up to 8-10 plates per week employ anywhere from 4-9 microplate positions. This may sound like relatively many, however several tip racks of different sizes, a waste position for tips, sample and reagent plates, a destination plate, and even intermediate plates consume this space quickly. Larger platforms accommodate over 50 SBS format items on deck and provide options for stacking in high-throughput screening experiments. As liquid handling products accommodate increasing throughputs, they usually increase in both size and price.



The ability to process plates more quickly comes from an ability to parallelize sample processing through the use of higher-channel pipetting heads (96/384), but also from the use of multiple pipetting heads. From systems slowly processing samples using a single channel head through systems quickly processing plates at a time by running 384-channel and 8-channel heads in parallel, researchers have a wide range of sample processing speeds to select from. Important considerations include unnecessary tip consumption associated with parallel processing and the expense of purchasing and maintaining costly 96- and 384- channel pipetting heads that require routine maintenance.


Pipetting Specifications and Reproducibility

Liquid handling manufacturers employ various techniques to achieve both liquid handling and supplemental sample handling steps of a given workflow. Interestingly, the technology employed for pipetting greatly impacts the accuracy and precision of liquid transfer volumes across varying liquid classes. In most cases, syringe-based pipetting of liquid or air will not provide the same level of reproducibility as piston-operated pipettes. Recently, products employing more traditional piston-operated pipetting, similar to the technology found inside manual pipettes, and positive displacement pipetting have entered the market to provide more reproducible liquid handling, especially at lower volumes. Keep an eye out for misleading specifications, and we advise to always request a fluorescence-based assessment of volumetric performance for each common sample and reagent utilized in your workflow. When it comes to full workflow automation of a manual process, check with liquid handling vendors to ensure that they can provide on-deck accessories for vital steps such as heating, cooling, mixing, shaking, centrifuging, extractions, and incubation.


Tip Consumption

Although the capital investment associated with an automated liquid handler can be daunting, also consider the cost of ownership associated with tip consumption and service when making a purchasing decision. Tip costs vary largely from vendor to vendor, and some platforms accommodate third party tips. Always inspect third party tips under a microscope to visualize any superfluous or rough plastic that may impact pipetting performance.



Liquid handling software either supports general liquid handling steps or a specific application. For common applications such as qPCR and Next-Generation Sequencing (NGS), software that integrates the components necessary for the entire workflow can be simplified so that users are not burdened with the complexity required to program individual liquid handling steps. However, if workflows are always changing in a given lab, and the instrument will be used for many differing application workflows, general purpose liquid handling software may make sense. Many liquid handling software packages require expert users to run, so make sure to participate in a software demonstration prior to purchasing to make sure that it fits well within the competencies of the staff in your lab.



Semi-Automated or “Hybrid” Liquid Handling

Currently, no liquid handler on the market features the perfect combination of cost-effective pipetting, reproducibility, ease-of-use, throughput, and speed. For instance, higher throughput workstations process hundreds of samples quickly, but pipette tips are costly, pipetting performance leaves much to be desired, and the software is almost impossible to program without some software development experience. On the other side of the spectrum, smaller application specific instruments offer wizard based software packages that make setting up routine qPCR experiments simple. Further, the pipetting performance matches that of electronic pipettes, but the instruments are slow, the throughput is limited, and the lack of accessories limits the degree of automation that can be obtained.


Some vendors have found a reasonably nice market solution that offers quite usable software accommodating full application workflows such as NGS, but like the rest of the market offerings, the unnecessary tip consumption for reagent dispensing and the air displacement pipetting leaves something to be desired across varying liquid classes.


A hybrid liquid handling approach allows researchers to leverage novel systems and technologies alongside one another to automate a small or specific part of a given workflow. By purchasing smaller liquid handling products that have been developed with a specific workflow step and customer benefit in mind, users can develop a liquid handling approach that leverages the strengths of many technologies and platforms allowing for the utilization of positive displacement pipetting and the conservation of tips and reagents, for instance.


The FORMULATRIX® MANTIS® is the cornerstone of any efficient and reproducible liquid handling arsenal. The MANTIS dispenses reagents across 384-well plates in minutes, allowing for a single tip to be consumed with no cross-contamination, and coupled with sample pipetting, researchers can achieve miniaturized reaction volumes reproducibly for the most cost-effective genomics experimentation possible today. Without the software requirements of traditional liquid handlers, the MANTIS coupled with a plate stamping instrument, for instance, allows for a high degree of both efficiency and process safety in genomics and drug discovery research.

The following is an example of a cost-effective, efficient, and reproducible liquid handling workflow for qPCR plate preparation from purified nucleic acids through complete qPCR plate preparation:

  1. Master mix preparation in 1.5 mL tubes with electronic single channel pipette*
  2. Sample transfer and serial dilution with electronic 8-channel pipette*
  3. Sample transfer and aliquot into triplicate reactions with 8-channel pipette*
  4. Master mix and primer addition across entire plate with the MANTIS Liquid Handler

*Alternatively, for a higher degree of automation, an entry level 8-channel liquid handling workstation can replace the single and multi-channel pipettes for master mix preparation and serial dilution respectively.