Quantitative multiplexed simulated-cell identification by SERS in microfluidic devices.

A reliable identification of cells on the basis of their surface markers is of great interest for diagnostic and therapeutic applications. We present a multiplexed labeling and detection strategy that is applied to four microparticle populations, each mimicking cellular or bacterial samples with varying surface concentrations of up to four epitopes, using four distinct biotags that are meant to be used in conjunction with surface enhanced Raman spectroscopy (SERS) instead of fluorescence, together with microfluidics. Four populations of 6 µm polystyrene beads were incubated with different mixtures, "cocktails" of four SERS biotags (SBTs), simulating the approach that one would follow when seeking to identify multiple biomarkers encountered in biological applications. Populations were flowed in a microfluidic flow-focusing device and the SERS signal from individual beads was acquired during continuous flow. The spectrally rich SERS spectra enabled us to separate confidently the populations by utilizing principal component analysis (PCA). Also, using classical least squares (CLS), we were able to calculate the contributions of each SBT to the overall signal in each of the populations, and showed that the relative SBT contributions are consistent with the nominal percentage of each marker originally designed into that bead population, by functionalizing it with a given SBT cocktail. Our results demonstrate the multiplexing capability of SBTs in potential applications such as immunophenotyping.

An injection molded microchip for nucleic acid purification from 25 microliter samples using isotachophoresis.

We present a novel microchip device for purification of nucleic acids from 25µL biological samples using isotachophoresis (ITP). The device design incorporates a custom capillary barrier structure to facilitate robust sample loading. The chip uses a 2mm channel width and 0.15mm depth to reduce processing time, mitigate Joule heating, and achieve high extraction efficiency. To reduce pH changes in the device due to electrolysis, we incorporated a buffering reservoir physically separated from the sample output reservoir. To reduce dispersion of the ITP-focused zone, we used optimized turn geometries. The chip was fabricated by injection molding PMMA and COC plastics through a commercial microfluidic foundry. The extraction efficiency of nucleic acids from the device was measured using fluorescent quantification, and an average recovery efficiency of 81% was achieved for nucleic acid masses between 250pg and 250ng. The devices were also used to purify DNA from whole blood, and the extracted DNA was amplified using qPCR to show the PCR compatibility of the purified sample.