Bioanalysis in Femtoliter Scale Printed Artificial Systems

Abstract

Printing has fueled the development of a new class of artificial biosystems for the qualitative and quantitative determination of bioanalytes [1]. In particular, the multiscale organization (from nanometers to millimeters) and multiplexed molecular composition (DNA, proteins, lipids, polymers) of such platforms enable the determination of molecular interactions in conditions mimicking/redesigning those of the living systems. Since the development of microarrays platforms [2], the downsizing of the “analyzable” feature down to femtoliter (fL) scales has permitted to develop new researches in the field of molecular condensates/confinement. Herein, two relevant examples of fL-scale systems will be discussed, highlighting their applications in bioanalysis. The first one includes molecular inks containing DNA nanoswitches or CYP2E1 catalyzed enzymatic reactions produced by inkjet printing fL-scale compartments into mineral oil drops [3]. The downscaling triggers the organization of a confined environment at the water/oil interface, resulting in up-concentration effects and molecular crowding. The employ of fluorescence lifetime imaging permits to analyze downscaling induced effects, namely up-concentration, heterogeneity and molecular proximity. The second example consists in fL-scale droplets produced by microchannel cantilever spotting (μCS) of inks containing single strand DNA (ssDNA) into porous substrates (nylon), resulting in oligonucleotides microarrays for sensing applications [4]. The downscaling to fL-scale imbibition into porous substrates highlights an intriguing and complex process controlled by the interplay of spreading, evaporation and capillary effects, being facilitated by glycerol additive in the ink. The DNA sequences functionality is demonstrated by hybridization with a fluorolabeled complementary sequence, producing a double strand sequence (dsDNA). The signal distribution in the spot is homogeneous and allows for the optical detection of spotted oligonucleotides down to few tents of zeptomoles.