Size‐based proteins separation using polymer‐entrapped colloidal self‐assembled nanoparticles on‐chip

We report on a facile method to stabilize colloidal self‐assembled (CSA) nanoparticles packed in microchannels for high speed size‐based separation of proteins. Silica nanoparticles, self‐assembled in a network of microfluidic channels, were stabilized with a methacrylate polymer prepared in situ through photopolymerization. The entrapment conditions were investigated to minimize the effect of the polymer matrix on the structure of the packing and the separation properties of the CSA beds. SEM shows that the methacrylate matrix links the nanoparticles at specific sphere–sphere contact points, improving the stability of the CSA structure at high electric fields (up to at least 1800 V/cm), allowing fast and efficient separation. The %RSD of the protein migration times varied between 0.3 and 0.5% (n = 4, in 1 day) and <0.83% over a period of 7 days (n = 28 runs) in a single device, at high field strength. The overall %RSD of protein migration times from chip‐to‐chip across a single fabrication run was 4.3% (n = 3) and between fabrication runs was 11% (n = 35), with 87% fabrication yield, demonstrating reproducible packing and entrapment behavior. The optimized entrapped CSA beds demonstrated better separation performance (plate height, H ～ 200 nm) than similarly prepared on‐chip CSA beds without the polymer entrapment. Polymer‐entrapped CSA beds also exhibited superior protein resolving power: the minimum resolvable molecular weight difference of proteins in the polymer‐entrapped CSA bed is 0.6 kDa versus ～9 kDa for the native silica CSA bed (i.e. without polymer entrapment).