Recent Advances in High-throughput Platforms with Engineered Biomaterial Microarrays for Screening of Cell and Tissue Behavior

Background

Cell adhesion, proliferation and differentiation depend on biochemical cues such as extracellular matrix (ECM) composition and soluble factors, and on biophysical cues such as topography, stiffness and applied mechanical or electrical inputs. Conventional in vitro screening is limited in accuracy, reusability, labor and versatility, and cannot cover the large number of combinations of these factors. High-throughput screening uses small amounts of material and cells and tests many conditions on one device, but 2D microarrays do not represent the 3D environment of cells in the body, and continuous monitoring of cell behavior over time requires integrated biosensors.

How the review is organized

The review is organized into three approaches. The first is screening of biochemical microenvironments with 2D and 3D biomaterial microarrays, made by contact printing, soft lithography or superhydrophobic-hydrophilic wettability patterning, and with microfluidic gradient generators. The second is screening of biophysical microenvironments, covering micro- and nanotopography, substrate stiffness and interfacial geometry, and active mechanical and electrical stimulation. The third is integration of optical and electrochemical biosensors with microfluidic and organ-on-a-chip platforms for continuous monitoring.

Scope of the review

For each approach the review describes representative platforms, the cells or tissues tested and the reported readouts. Examples include a polymer microarray of 496 compositions from 22 acrylate monomers for human embryonic stem cell (hESC) self-renewal, a Topochip of 2,176 topographies, polyacrylamide stiffness arrays in 96- and 384-well plates, compression and magnetic stretching arrays for cell-laden hydrogels, and sensors for pH, oxygen and secreted biomarkers such as albumin, glutathione S-transferase alpha (GST-α) and creatine kinase MB (CK-MB). Figure 1 to Figure 5 reproduce panels from the cited studies.

Main points of the review

For biochemical screening, the review describes contact-printed ECM and polymer arrays, in which high-acrylate polymers with moderate wettability enhanced hESC colony formation, and wettability-patterned arrays such as one that tested fibroblasts and pre-osteoblasts in 24 hydrogel compositions. Microfluidic platforms generated spatial chemical gradients, for example interleukin-1β from 0.005 to 0.1 ng/mL across a culture chamber, and gradient-fed hanging-drop arrays followed spheroid growth over 60 hours.

For biophysical screening, a T cell platform showed increased interleukin-2 production on trench-grid surfaces with 200 nm trench width and grating pitch above 2.3 μm, and lower production at 100 nm width and pitch below 0.5 μm. Polyacrylamide arrays spanned 0.3 to 55 kPa and were used with 7 cell types, and in micropatterned hydrogels of 11 shapes, spiral shapes gave the highest cancer stem cell marker expression while matrix stiffness had no significant effect. Mechanical platforms applied compressive strain from 0% to 26% or magnetic strain from 0 to 60%, and the review notes that these platforms addressed only a few combinations of mechanical input; Seo and colleagues’ interconnectable bioreactors, with gas-actuated microposts, were described as screening GelMA concentration and dynamic compression together. Electrical stimulation examples included a triboelectric generator delivering pulses of 1 Hz and about 270 nA and flexible nanoelectronics that manipulated action potential propagation in 3D cardiac tissue with stimuli of 1 V and 1.25 Hz.

For sensor integration, the review describes an optical module that read pH from phenol red absorbance and oxygen from fluorophore quenching, a bead-based electrochemical immunosensor with a detection limit of 0.03 ng/mL against 0.2 ng/mL for ELISA, and an aptamer sensor that detected CK-MB from a heart-on-a-chip exposed to doxorubicin. Regenerable electrochemical sensors restored electrode current to close to 100% of the as-fabricated level, and a multisensor liver-and-heart platform monitored albumin, GST-α and CK-MB during drug exposure for up to 5 days. Droplet microarrays with individually addressable electrodes for up to 20 electrochemical cells and a single-droplet colorimetric assay that moves droplets by vacuum-driven local deformation of a patterned PDMS substrate were presented as routes toward high-throughput sensing.

Open questions

The review states that simultaneous screening and analysis of cellular behavior remains a challenge and calls for standardized platforms compatible with common culture and analysis methods and capable of in situ monitoring. It notes that regenerated electrodes last only a finite number of cycles, that PDMS in organ-on-a-chip devices can adsorb and absorb drugs, and that the multisensor platform is large and complex. A microfluidics-integrated high-throughput biosensor had not yet been achieved, and screening under dynamic microenvironments with such sensors remained unexplored.

How it connects to other research

Mei and colleagues, cited in this review, screened synthetic polymer substrates for human pluripotent stem cell culture and related wettability, topography, surface chemistry and modulus to colony formation; the review places this kind of 2D materials screening alongside 3D, biophysical and sensor-integrated platforms.