Biomarker barcodes: multiplexed microfluidic immunohistochemistry enables high-throughput analysis of tissue microarray.

We present a multiplexed microfluidic immunohistochemistry (IHC) technology that enables high-throughput analysis of tissue microarrays (TMAs) using the patterns of biomarker barcodes, which consist of a series of expressed linear patterns of specific biomarkers. A multichannel poly(dimethylsiloxane) microfluidic device was reversibly assembled by the pressure of simple equipment for multiplexed IHC on each core of TMA or cell microarray (CMA) section slides. By injecting primary antibodies from different biomarkers independently into each channel, multiplexed immunostaining can be performed on each core of TMA. We confirmed the equal immunostaining quality regardless of the channel orders and core positions in the slide. Four different biomarkers (ER, PR, HER2, and Ki67) were used for the demonstration of distinctive expression patterns on CMAs which consist of six different breast cancer cell lines, and it was confirmed that these bar-like signals could be a biomarker barcode for the TMA core. A biomarker barcode of breast cancer patient-derived TMA was quickly scanned by a slide scanner and compared to the conventional method for breast cancer diagnosis. This "barcode-IHC" concept, which has been verified by performing multiplexed microfluidic IHC on CMA and TMA samples, provides high reproducibility and the potential of high-throughput screening with molecular diagnostic capability.