Effect of design geometry, exposure energy, cytophilic molecules, cell type and load in fabrication of single-cell arrays using micro-contact printing

In this study a range of factors influencing the fabrication of single-cell arrays (SCAs) are identified and investigated. Micro-contact printing was used to introduce spots coated with polyethyleneimine or Matrigel on glass surfaces pre-coated with polyethylene glycol. Unmodified E. coli, Synechococcus sp., Chlamydomonas reinhardtii as well as diverse mammalian cells including HUVEC, AAV293, U87, OHS, PC3, SW480, HepG2 and AY-27 were successfully immobilised onto the chemically coated spots. The developed SCAs show high cell viability and probability for capturing single-cells. A discrepancy between the size and shape of the squares described in the design file and the actual structures obtained in the final PDMS structure is characterised and quantified. The discrepancy is found to be depending on the exposure energy used in the photolithography process as well as the size of the squares and their separation distance as detailed in the design file. In addition to these factors, the effect of the cell density loaded onto the patterned surfaces is also characterised. The systematic characterisation of key parameters that need to be optimised prior to the fabrication of SCAs is essential in order to increase the efficiency and reproducibility of future fabrication of SCAs for single-cell studies.