Abstract
In vitro assays are an important platform for cancer research as they allow high-throughput experimentation that is not possible using in vivo animals. Although various in vitro assays are developed to study cell viability or migration, many of these assays are often limited to two dimensions, involving complex procedures or relying specialized equipment, etc. Here, we designed a simple colorimetric assay that accommodates automatic liquid samples loading, high-throughput generation of chemical concentration gradient, three-dimensional (3D) cell culture establishment, and smartphone-based colorimetric readouts. This assay is based on through-hole arrays in the poly(methyl methacrylate) (PMMA) layers. Liquid samples can be automatically loaded into through-hole arrays in PMMA layers by capillary force. Different drug concentrations can be generated by aligning and stacking to mix the contents of the corresponding through-holes with different volumes. 3D culture of cancer cells can be established by the rapid absorption of cell suspensions into the macroporous gels. After exposing the 3D cultured cells to different drug concentrations, the number of viable cells and migrated cells was reflected by the color change of Alamar blue, which enable on-site readout by a smartphone. This assay can study cell viability as well as cell migration, the two main characteristics of cancer cells, using one device. Interestingly, HeLa cells remained with high viability after cryopreservation at -80 °C, which allows for storage and distribution using dry ice. The simple protocol, along with the cryopreservability at -80 °C facilitates its ease of use to study cell viability together with cell migration in common laboratories or clinical settings.