Detecting normal and cancer skin cells via glycosylation and adhesion signatures: A path to enhanced microfluidic phenotyping.

Recruiting circulating cells based on interactions between surface receptors and corresponding ligands holds promise for capturing cells with specific adhesive properties. Our study investigates the adhesion of skin cells to specific lectins, particularly focusing on advancements in lectin-based biosensors with diagnostic potential. We explore whether we can successfully capture normal skin (melanocytes and keratinocytes) and melanoma (WM35, WM115, WM266-4) cells in a low-shear flow environment by coating surfaces with lectins. Specifically, we coated surfaces with Dolichos biflorus (DBA) and Maackia Amurensis (MAL) lectins, which were used to detect and capture specific skin cells from the flow of cell mixture. Alterations in glycan expression (confirmed by fluorescent microscopy) demonstrated that DBA binds predominantly to normal skin cells, while MAL interacts strongly with melanoma cells. Assessing adhesion under static and dynamic low-shear stress conditions (up to 30 mPa) underscores the reliability of DBA and MAL as markers for discriminating specific cell type. Melanocytes and keratinocytes adhere to DBA-coated surfaces, while melanoma cells prefer MAL-coated surfaces. A comprehensive analysis encompassing cell shape, cytoskeleton, and focal adhesions shows the independence of our approach from the inherent characteristics of cells, thus demonstrating its robustness. Our results carry practical implications for lectin-biosensor designs, emphasizing the significance of glycan-based discrimination of pathologically altered cells. Combined with microfluidics, it demonstrates the value of cell adhesion as a discriminant of cancer-related changes, with potential applications spanning diagnostics, therapeutic interventions, and advanced biomedical technologies.

Plasma Treatment of PDMS for Microcontact Printing (µCP) of Lectins Decreases Silicone Transfer and Increases the Adhesion of Bladder Cancer Cells.

The present study investigates silicone transfer occurring during microcontact printing (µCP) of lectins with polydimethylsiloxane (PDMS) stamps and its impact on the adhesion of cells. Static adhesion assays and single-cell force spectroscopy (SCFS) are used to compare adhesion of nonmalignant (HCV29) and cancer (HT1376) bladder cells, respectively, to high-affinity lectin layers (PHA-L and WGA, respectively) prepared by physical adsorption and µCP. The chemical composition of the µCP lectin patterns was monitored by time-of-flight secondary ion mass spectrometry (ToF-SIMS). We show that the amount of transferred silicone in the µCP process depends on the preprocessing of the PDMS stamps. It is revealed that silicone contamination within the patterned lectin layers inhibits the adhesion of bladder cells, and the work of adhesion is lower for µCP lectins than for drop-cast lectins. The binding capacity of microcontact printed lectins was larger when the PDMS stamps were treated with UV ozone plasma as compared to sonication in ethanol and deionized water. ToF-SIMS data show that ozone-based treatment of PDMS stamps used for µCP of lectin reduces the silicone contamination in the imprinting protocol regardless of stamp geometry (flat vs microstructured). The role of other possible contributors, such as the lectin conformation and organization of lectin layers, is also discussed.

Bladder Cancer Cells Interaction with Lectin-Coated Surfaces under Static and Flow Conditions.

Aberrant expression of glycans, i.e., oligosaccharide moiety covalently attached to proteins or lipids, is characteristic of various cancers, including urothelial ones. The binding of lectins to glycans is classified as molecular recognition, which makes lectins a strong tool for understanding their role in developing diseases. Here, we present a quantitative approach to tracing glycan-lectin interactions in cells, from the initial to the steady phase of adhesion. The cell adhesion was measured between urothelial cell lines (non-malignant HCV29 and carcinoma HT1376 and T24 cells) and lectin-coated surfaces. Depending on the timescale, single-cell force spectroscopy, and adhesion assays conducted in static and flow conditions were applied. The obtained results reveal that the adhesion of urothelial cells to two specific lectins, i.e., phytohemagglutinin-L and wheat germ agglutinin, was specific and selective. Thus, these lectins can be applied to selectively capture, identify, and differentiate between cancer types in a label-free manner. These results open up the possibility of designing lectin-based biosensors for diagnostic or prognostic purposes and developing strategies for drug delivery that could target cancer-associated glycans.

Correction: A system for the high-throughput measurement of the shear modulus distribution of human red blood cells.

Correction for 'A system for the high-throughput measurement of the shear modulus distribution of human red blood cells' by Amir Saadat et al., Lab Chip, 2020, 20, 2927-2936, DOI: 10.1039/D0LC00283F.

A system for the high-throughput measurement of the shear modulus distribution of human red blood cells.

Reduced deformability of red blood cells (RBCs) can affect the hemodynamics of the microcirculation and reduce oxygen transport efficiency. It is also well known that reduced RBC deformability is a signature of various physical disorders, including sepsis, and that the primary determinant of RBC deformability is the membrane shear modulus. To measure the distribution of an individual's RBC shear modulus with high throughput, we a) developed a high-fidelity computational model of RBCs in confined microchannels to inform design decisions; b) created a novel experimental system combining microfluidic flow, imaging, and image analysis; and c) performed automated comparisons between measured quantities and numerical predictions to extract quantitative measures of the RBC shear modulus for each of thousands of cells. We applied our computational simulation platform to construct the appropriate deformability figure(s) of merit to quantify RBC stiffness based on an experimentally measured, steady-state cell shape in flow through a microchannel. In particular, we determined a shape parameter based on the second moment of the cell shape that is sensitive to the changes in the membrane stiffness and cell size. We then conducted microfluidic experiments and developed custom automated image processing codes to identify and track the position and shape of individual RBCs within micro-constrictions. The fabricated microchannels include a square cross-section imaging region (7 by 7 µm) and we applied order 10 kPa pressure differences to induce order 10 mm s-1 cell velocities. The combination of modeling, microfluidics, and imaging enables, for the first time, quantitative measurement of the shear moduli of thousands of RBCs in human blood samples. We demonstrate the high-throughput features by sensitive quantification of the changes in the distribution of RBC stiffness with aging. This combined measurement and computational platform is ultimately intended to diagnose blood cell disorders in patients.