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Analysis in Cuvette Holder Conserves Samples

Wed, 12/07/2011 - 9:32am
Jonathan Redfern, General Manager; Picodrop Ltd., Saffron Walden, U.K.
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Spectrophotometers are among the many analytical tools found on every scientist’s bench top. By passing light of a known frequency through a sample and measuring what is absorbed, the instrument yields information about the contents of the sample. DNA, RNA, proteins—in short, most materials that are used in today’s biological research and development—can all be measured using a spectrophotometer.

Most molecular biology assays require an initial assessment of sample concentration and purity before subsequent dilutions and downstream processing can commence. Traditionally, samples have been measured in cuvettes requiring significant dilution of the sample before sufficient volume can be produced to allow measurement in a 10 mm cuvette (typically up to 2 mL). The dilution of samples leads to inaccuracy of results, and means the sample cannot be reused.

Instruments from different manufacturers—designed to quantify small volumes without dilution—use similar lamp and fiber optics and have similar technical specifications for minimum concentrations of approximately 3 ng/µL +/-2 ng DNA.

The Picodrop P200 has an interchangeable cuvette holder that can be used for detection of dilute samples down to 0.5 ng/µL +/-0.2 ng. A dual circuit optical loop continuously measures the lamp drift and deducts any variation from the sample background absorbance.

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The other systems use a method of dropping the sample directly on to a sample platform or pedestal and then closing a lid over the sample. Thorough cleaning of both surfaces is then required before the next sample can be added. Samples cannot be recovered easily.

The Picodrop system uses a pipette tip as the sample container. The sample is contained within the tip at all times and can be recovered by pipeting out from the same tip, reducing sample usage and potential cross contamination

The design of Picodrop’s fiber optics and ball lenses combined with the availability of thermostable and optically clear plastics, make in-tip detection a reality. Ultraviolet (UV) transparent polymers were used to developed a UV-transparent pipette tip to replace the standard cuvette.

UV light is passed directly through the bottom 1 mm of the tip and UV/Vis absorbance of its contents are calculated. This design allows volumes down to 2 µL to be used.

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