The pipetting of serial dilution series is daily business in biomedical laboratories, be it for the determination of the effect of test compounds in biochemical or cellular assays, for finding optimal concentrations of substrates and co-factors in assay development, or for a wide range of enzymological studies, such as active site titrations or determinations of kinetic constants of slow binding and irreversible inhibitors.
In this article, we introduce a benchtop instrument, HP D300 Digital Dispenser, available from Tecan, that reliably and rapidly dispenses small volumes of DMSO-dissolved compounds directly into wells of microtiter plates in a contact-free manner directly from stock solutions. The device dispenses single droplets of either 20 or 13 picoliters, and larger volumes are digitally built by rapidly shooting multiple droplets into the same well, just like ink-jet printers shoot ink droplets on paper to print letters and words.
The remarkably small volume that is transferred with a single shot allows the direct dispensing of stock solutions into assay plates, spanning a wide range of concentrations of the dispensed compound. It is possible to generate custom-made dilution series of DMSO-dissolved compounds rapidly and accurately, with minimized compound consumption and with minimal effort.
Operating the instrument is simple. First, parameters such as the plate format (12- to 384-well plates are supported), the assay volume, the maximally tolerated DMSO concentration, and the names and stock concentrations of the compounds to be dispensed are defined. The simplest way to subsequently program a dilution series is to highlight a certain number of wells, and then define the desired starting and end concentrations of the compound and decide whether the dilution scheme should be logarithmic or linear.
When starting the dispensing procedure, the program asks the user to pipet the compound into a dispensehead reservoir (usually a few microliters), and the instrument then shoots the compound directly into the wells of the microtiter plate as programmed. Completely custom-made dilution series or complicated matrix experiments can thus be prepared in microtiter plates in a few minutes with minimal manual work.
In this tutorial, we show two different applications for the use of digital dispensing, namely the determination of kinetic parameters of enzyme-catalyzed reactions and the direct dispensing of test compounds from a DMSO stock solution onto living cells.