  We present a new methodology for efficient and high-quality patterning of
biological reagents for surface-based biological assays. The method relies on
hydrodynamically confined nanoliter volumes of reagents to interact with the
substrate at the micrometer-length scale. We study the interplay between
diffusion, advection, and surface chemistry and present the design of a
noncontact scanning microfluidic device to efficiently present reagents on
surfaces. By leveraging convective flows, recirculation, and mixing of a
processing liquid, this device overcomes limitations of existing biopatterning
approaches, such as passive diffusion of analytes, uncontrolled wetting, and
drying artifacts. We demonstrate the deposition of analytes, showing a 2- to
5-fold increase in deposition rate together with a 10-fold reduction in analyte
consumption while ensuring less than 6% variation in pattern homogeneity on a
standard biological substrate. In addition, we demonstrate the recirculation of
a processing liquid using a microfluidic probe (MFP) in the context of a
surface assay for (i) probing 12 independent areas with a single microliter of
processing liquid and (ii) processing a 2 mm2 surface to create 170 antibody
spots of 50 x 100 {\mu}m2 area using 1.6 {\mu}L of liquid. We observe high
pattern quality, conservative usage of reagents, micrometer precision of
localization and convection-enhanced fast deposition. Such a device and method
may facilitate quantitative biological assays and spur the development of the
next generation of protein microarrays.

Biomolecules