Deep phenotyping of cancer cells at the single-cell level is of critical importance in the era of precision medicine to advance understanding of the precise relationship between gene mutation and cell phenotype and to elucidate biological nature of tumor heterogeneity and their potential biological and clinical implications. Existing microfluidic single-cell phenotyping tools, albeit their high-throughput, high-resolution operation, are limited to phenotypic measurements of 1 – 2 selected morphological and physiological features of single cells. To address the critical need for multiplexed, informative phenotyping of live single cancer cells, herein we reported a microfluidic elasticity microcytometer for multiparametric biomechanical and biochemical phenotypic profiling of free-floating, live single cancer cells to obtain quantitative information of cell size, cell deformability / stiffness, and expression levels of surface receptors simultaneously for the same single live cancer cells. The elasticity microcytometer was implemented for single-cell measurements and comparisons of four human cell lines with distinct metastatic potentials and derived from different human tissues. An analytical model was developed from first principles for the first time to effectively convert cell deformation and adhesion information of single cancer cells encapsulated inside the elasticity microcytometer to cell deformability / stiffness and surface protein expression. Together, the elasticity microcytometer holds a great promise for comprehensive molecular, cellular, and biomechanical phenotypic profiling of live cancer cells at the single cell level, critical for studying intra-tumor cellular and molecular heterogeneity using low-abundance, clinically relevant human cancer cells.

Transendothelial migration of cancer cells is a critical stage in cancer, including breast cancer, as the migrating cells are generally believed to be highly metastatic. However, it is still challenging for many existing platforms to achieve a fully covering endothelium and to ensure transendothelial migration capability of the extracted cancer cells for analyses with high specificity. Here, we report a microfluidic device containing multiple independent cell collection microchambers underneath an embedded endothelium such that the transendothelial-migrated cells can be selectively collected from only the microchambers with full coverage of an endothelial layer. In this work, we first optimize the pore size of a microfabricated supporting membrane for the endothelium formation. We quantify transendothelial migration rates of a malignant human breast cell type (MDA-MB-231) under different shear stress levels. We investigate characteristics of the migrating cells including morphology, cytoskeletal structures, and migration (speed and persistence). Further implementation of this endothelium-embedded microfluidic device can provide important insights into migration and intracellular characteristics related to cancer metastasis and strategies for effective cancer therapy.

Microcontact printing (μCP) is widely used to create patterns of biomolecules essential for studies of cell mechanics, migration, and tissue engineering. However, different types of μCPs may create micropatterns with varied protein-substrate adhesion, which may change cell behaviors and pose uncertainty in result interpretation. Here, we characterize two μCP methods for coating extracellular matrix (ECM) proteins (stamp-off and covalent bond) and demonstrate for the first time the important role of protein-substrate adhesion in determining cell behavior. We found that, as compared to cells with weaker traction force (e.g., endothelial cells), cells with strong traction force (e.g., vascular smooth muscle cells) may delaminate the ECM patterns, which reduced cell viability as a result. Importantly, such ECM delamination was observed on patterns by stamp-off but not on the patterns by covalent bonds. Further comparisons of the displacement of the ECM patterns between the normal VSMCs and the force-reduced VSMCs suggested that the cell traction force plays an essential role in this ECM delamination. Together, our results indicated that μCPs with insufficient adhesion may lead to ECM delamination and cause cell death, providing new insight for micropatterning in cell-biomaterial interaction on biointerfaces.

Phenotypic profiling of single floating cells in liquid biopsies is the key to the era of precision medicine. Random laser in biofluids is a promising tool for the label-free characterization of the biophysical properties due to the high brightness and sharp peaks of the lasing spectra, yet previous reports were limited to the random laser in solid tissues with dense scatterings. In this report, a random laser cytometer is demonstrated in an optofluidic device filled with gain medium and human breast normal/cancerous cells. The multiple light scattering induced by the micro-scale human cells promotes the random lasing and influences the lasing properties in term of laser modes, spectral wavelengths, and lasing thresholds. A sensing strategy based on analyzing the lasing properties to determine both the whole-cell and the subcellular biophysical properties of the cells and the malignant alterations of the cell suspensions are successfully detected. Our results provide a new approach to designing a label-free biophysical cytometer based on optofluidic random laser devices, which is advantageous for further research in the field of random laser bio-application.