Artel finds Death Valley's heat devastates accurate pipetting
US, to see the size of errors that dry heat would introduce to
pipetted volumes and found they could be as high as 35 per cent.
Laboratory workers commonly encounter temperature, pressure and humidity changes in the laboratory and Artel's "Extreme Pipetting" expeditions have been designed to highlight the impact that these environmental factors can have on the accuracy and precision of pipettes. The latest expedition went to Badwater in Death Valley, one of the hottest and driest places in the world where temperatures can commonly exceed 49°C (120°F) as well as having a relative humidity of around 6 per cent. Badwater gets its name from a small spring-fed pool that is so full of salts that have accumulated due to water evaporation that it is undrinkable. The team, led by Dr George Rodrigues, senior scientific manager at Artel, investigated the effect that the extreme conditions in Death Valley had on pipetted volumes. "We want lab technicians to think about their local environment, the temperature, humidity and barometric pressure and how that might affect their data," said Dr Rodrigues. Inaccurate pipetting can lead to large errors and cause expensive QC (quality control) issues and problems with regulatory bodies if the errors remain undetected until after products have been released. To counter this issue, Artel has developed a range of instruments that enables liquid handling devices, and their operators, to be calibrated in the laboratory that they are used in rather than in the factory preset conditions. These instruments use a ratiometric photometry technique and include the PCS (pipette calibration system) for calibrating single-channel pipettes and the MVS (multichannel verification system) for calibrating automated high-throughput liquid handling devices. "Many pharmaceutical QC labs undergo temperature changes of up to 5°C during the day, and if the humidity is not controlled this can lead to nearly a 20 percentage point change in the relative humidity and introduce errors in liquid dispensing," said Dr Rodrigues. Artel's first expedition to highlight how environmental conditions affect pipetting accuracy took the team to the top of Mount Washington, US, where the pressure is so low that water boils at 93°C and induces a 9 per cent error when pipetting from a 1µl pipette. To ensure that the pipetting was consistent and that the errors measured were due to the environment and not the user, the team were all trained in the Artel pipetting method. The second expedition saw the team travel to Yellowstone National Park, US, where errors of up to 37 per cent were observed when pipetting cold liquids (3°C) and up to 24 per cent when the liquids were warm (37°C). Geoff Sawyer, a member of the Extreme Pipetting Expedition, likened the heat the team experienced during this latest expedition to Death Valley to "the rush of heat you feel when opening an oven set on high heat." They found that using standard pipetting techniques a 2µl pipette set to deliver is minimum volume of 0.2µl under delivered by 34.7 percent, way over the 12 per cent minimum volume accuracy specified by the manufacturer. When set to its maximum volume of 2µl, the pipette under-delivered by 7 per cent. These errors could be reduced somewhat by "pre-wetting" the pipette tip with the errors dropping to 30.0 per cent for a 0.2µl delivery and 4.8 per cent for a 2µl delivery. These errors were lessened when working with larger liquid volumes, with a 20µl pipette under-delivering by 16.8 per cent when set to its minimum volume. Pre-wetting the pipette tip reduced this error to 7.4 per cent. Unlike the previous trips there is, as yet, no full proof method for eliminating these errors, although the data suggest that pipetting at maximum volume and pre-wetting the pipette tip significantly reduce errors occurring. "Based on this data, we highly recommend pre-wetting pipette tips to reduce volume loss when pipetting in dry and hot environments," said Doreen Rumery, laboratory technical manager and quality control manager at Artel. The company ascribe the errors to the evaporation of the liquid in the pipette, where even a miniscule amount of liquid evaporation can cause a large error - far more than is actually evaporated. This is because when 1µl of liquid evaporates it converts into more than 1000µl of gas, expanding by a factor of up to 1,450 depending on the range and preventing the pipette from aspirating the desired amount. Pre-wetting the pipette not only helps to cool the tip but also creates a damp environment in the pipette that somewhat reduces the propensity of the liquid to evaporate due to shifting the evaporation equilibrium point. "It would be difficult to calibrate all of the error out of these measurements if pipetting by hand as the heat causes the liquid to evaporate and increase the pressure in the pipette, forcing out droplets of liquid and causing the pipette to under deliver," said Dr Rodrigues. "It may be possible to counter this problem using an automated liquid handler that had a precise time between aspiration and delivery so that the system could be calibrated to the exact temperature and amount of time that the liquid had to evaporate."
