Frøslev, Tobias Guldberg

Hansen, Anders Johannes

Morling, Niels

Stangegaard, Michael

English

Copenhagen University Research Information System

A simple method for validation and verification of pipettes mounted on automated liquid handlers

u n i ve r s i t y  o f  co pe n h ag e n  Københavns UniversitetA simple method for validation and verification of pipettes mounted on automatedliquid handlersStangegaard, Michael; Hansen, Anders Johannes; Frøslev, Tobias Guldberg; Morling, NielsPublication date:2009Document versionPublisher's PDF, also known as Version of recordCitation for published version (APA):Stangegaard, M., Hansen, A. J., Frøslev, T. G., & Morling, N. (2009). A simple method for validation andverification of pipettes mounted on automated liquid handlers. Poster session presented at LabAutomation2009,Palm Springs 2009, United States.Download date: 02. Feb. 2020A simple method for validation and verification of pipettes mounted on automated liquid handlersStangegaard M, Hansen AJ, Frøslev TG, Morling NSection of Forensic Genetics, Department of Forensic Medicine, Faculty of Health Sciences, University of Copenhagen, 11 Frederik V’s Vej, DK-2100 Copenhagen, DenmarkAutomated liquid handlers (ALHs) are increasingly used to improve throughput, pipetting  accuracy and prevent occupational injuries to the technical staff due to intensive manual pipetting1.  Standardized procedures for calibration of standard piston operated pipettes exist and many   laboratories calibrate their pipettes with defined intervals2.  However, less standardized is the routine validation and verification of ALHs, though   suggestions to how it should be performed exist3.  Commercial solutions have also been introduced4. Using pre-defined acceptance criteria, each pipette on each ALH was then either approved or rejected. Rejected pipettes were either repaired or the volume deviation was compensated for by applying a calibration curve in the liquid handler software. We have implemented the method on a Sias Xantus, a MWGt TheONYX, four Tecan Freedom EVO 150, a Biomek NX Span-8 and four Biomek 3000 robots. A 7-step serial dilution of Orange G was prepared manually in quadruplicates in a flat bottom 96-well microtiter plate (BD Falcon) by means of calibrated pipettes (column 9-12). This was used as a standard row. Each pipette of the liquid handler (1 up to 8) dispensed a selected volume (1 to 200µL) of Orange G eight or more times into the wells of the microtiter plate. All wells contained a total of 200µL liquid. The optical density (OD) was read at 490 nm, and the dispensed volume of each pipette was calculated based on a plot of volume and OD of a known set of Orange G dilutions. Finally, the percent inaccuracy (%d) and the imprecision (%CV) of each pipette was calculated.[1]  Bradshaw, J.T. et al., Multichannel  Verification  System (MVS): a Dual-Dye  Ratiometric  Photometry  system for performance verification  of multichannel  liquid  delivery  devices. JALA  10  (1), 35-42 (2005).[2]  Batista, E. et al., Volume calibration of 1000 μl  micropipettes. Inter-laboratory comparison. Accreditation and Quality Assurance: Journal for Quality, Comparability and Reliability in Chemical Measurement 13(4), 261-266 (2008).[3]  Taylor, P.B.  et al., A standard operating procedure for assessing liquid handler performance in high-  throughput screening. J Biomol  Screen  7  (6), 554-569 (2002).[4]  Bradshaw, J.T., et al., Multichannel Verification System (MVS): a Dual-Dye Ratiometric  Photometry system for performance verification of multichannel liquid delivery devices. JALA  10  (1), 35-42 (2005).  Figure 3. Two different volume verification runs both using 50µL as target volume on a 4 channel fixed tip MWG TheONYX  robot. Poster TP58LabAutomation2009Palm Springs, CA, USAMATERIAL AND METHODSINTRODUCTIONRESULTSCONCLUSIONSREFERENCESFigure 5. Three different volume verification runs all using 1.0µL as target volume on a Biomek NX span-8 using disposable tips. Run A and Run B show either too little or too large volume dispensed. Run C shows acceptable pipetting.Figure 4: Two different volume verification runs both using 5.0µL as target volume on an eight channel fixed tip Sias Xantus robot. Run A showed un-  acceptable pipetting. Run B following calibration showed acceptable   pipetting.Figure 1. Plate layout. A standard row is included on each plate.We have set up and implemented a simple solution for the continuous   verification of pipettes mounted on automated liquid handlers as  necessary for accredited work under the international laboratory standard ISO 17025. The method is cheap, simple and easy to use for aqueous solutions, but it requires a spectrophotometer that can read microtiter plates. The method can be used with both disposable tips, fixed tips as well as manual pipetting.  PROCESS FLOWThe process is composed of few and simple steps. The same stock solution may be used for verification of multiple ALHs. A calibrated ALH may be used to prepare the standard row. Initial calibration was performed with  manually pipetted  standard rows.Figure 2. Volume verification using manual pipetting  and a conventional pipette set to 2.0µL. Pipette number refers to different runs.

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A simple method for validation and verification of pipettes mounted on automated liquid handlers

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