Lateral Microscope Enables the Direct Observation of Cellular Interfaces and Quantification of Changes in Cell Morphology during Adhesion

Abstract

The ability to observe cell adhesion processes in real-time remains a grand challenge in basic biology and medicine. Toward this goal, we have developed a lateral optical microscope that allows direct observation of cell–substrate interactions in real-time on any substratetransparent, opaque, or coatedwithout requiring labels or specialized optical components. We demonstrate the use of our lateral microscope by quantifying dynamic changes in cell morphology during the first 90 min of adhesion to various materials. Specifically, we determined the rates of change in contact angle of HeLa, 3T3, HEK293, and MDA-MB-231 cells on five different substrates: glass, collagen-coated glass, Nylon, PTFE, and collagen-alginate hydrogels. We used these rates of change to compare adhesion of different cell lines on each surface, and to rank the adhesion-promoting capacities of the five surfaces for each cell line. For HeLa, 3T3, and HEK293 cells, we observed maximal rates of change in contact angle (0.058 min–1) on collagen-coated glass substrates. All five cell lines exhibited minimal rates of change (0.006 min–1) on PTFE. Lateral microscopy also revealed a unique morphology among MDA-MB-231 breast cancer cells during initial adhesion, which we quantified using measurements of changes in cell height. Our lateral microscope will not only enable more comprehensive, quantitative studies of cell adhesion to inform the development of biomaterials but will ultimately assist in advancing our understanding of many important biological processes and discovering new behaviors related to cell adhesion.

Effect of Functional Groups of Self-Assembled Monolayers on Protein Adsorption and Initial Cell Adhesion

“…Surface modifications using various physical, chemical, and biological processes had been reported to circumvent such unwanted processes. − Surfaces of ceramics, polymers, and metals have been modified via physical (for example, plasma treatment) and chemical methods (for example, functionalization via self-assembled monolayers [SAMs], immobilized ligands, etc.) to carefully modulate protein adsorption, cell adhesion, and differentiation. − …”

Correlation of Cell Surface Biomarker Expression Levels with Adhesion Contact Angle Measured by Lateral Microscopy

“…Such an approach has been applied successfully to studies of fundamental adhesion processes related to the differential adhesion hypothesis 24 and the differential interfacial tension hypothesis, 25 as well as to studies of adhesion interactions related to cell surface engineering and tissue engineering. 26,27 In this paper, we demonstrate the utility of lateral microscopy 28 to observe and quantify cell adhesion as it correlates with surface biomarker expression levels (Figure 1). We introduced different types of leukocytes to glass coverslips coated in surface antigen-specific antibodies (e.g., anti-CD4 specific for T cells expressing CD4).…”

“…In this paper, we demonstrate the utility of lateral microscopy to observe and quantify cell adhesion as it correlates with surface biomarker expression levels (Figure ). We introduced different types of leukocytes to glass coverslips coated in surface antigen-specific antibodies (e.g., anti-CD4 specific for T cells expressing CD4).…”

“…The Contact Angle plug-in for ImageJ was used to measure the contact angles formed between cells and antibody-coated substrates as previously described . Left and right contact angle measurements were generated for each cell, and the average of the two was reported and used to calculate the change in contact angle from 0 to 90 min.…”

“…The lateral microscope and sample containers to enable imaging in the lateral field of view were fabricated as previously described. 28 Images of leukocytes adhering to antibody-coated substrates were acquired as follows: (i) The live-cell enclosure surrounding the lateral microscope was allowed to warm to 37 °C for approximately 30 min. (ii) An antibody-coated substrate was added to an ethanolsterilized sample container and washed twice with RPMI.…”