Adhesion kinetics of human primary monocytes, dendritic cells, and macrophages: Dynamic cell adhesion measurements with a label-free optical biosensor and their comparison with end-point assays

Abstract

Monocytes, dendritic cells (DCs), and macrophages (MFs) are closely related immune cells that differ in their main functions. These specific functions are, to a considerable degree, determined by the differences in the adhesion behavior of the cells. To study the inherently and essentially dynamic aspects of the adhesion of monocytes, DCs, and MFs, dynamic cell adhesion assays were performed with a high-throughput label-free optical biosensor [Epic BenchTop (BT)] on surfaces coated with either fibrinogen (Fgn) or the biomimetic copolymer PLL-g-PEG-RGD. Cell adhesion profiles typically reached their maximum at ∼60 min after cell seeding, which was followed by a monotonic signal decrease, indicating gradually weakening cell adhesion. According to the biosensor response, cell types could be ordered by increasing adherence as monocytes, MFs, and DCs. Notably, all three cell types induced a larger biosensor signal on Fgn than on PLL-g-PEG-RGD. To interpret this result, the molecular layers were characterized by further exploiting the potentials of the biosensor: by measuring the adsorption signal induced during the surface coating procedure, the authors could estimate the surface density of adsorbed molecules and, thus, the number of binding sites potentially presented for the adhesion receptors. Surfaces coated with PLL-g-PEG-RGD presented less RGD sites, but was less efficient in promoting cell spreading than those coated with Fgn; hence, other binding sites in Fgn played a more decisive role in determining cell adherence. To support the cell adhesion data obtained with the biosensor, cell adherence on Fgn-coated surfaces 30–60 min after cell seeding was measured with three complementary techniques, i.e., with (1) a fluorescence-based classical adherence assay, (2) a shear flow chamber applying hydrodynamic shear stress to wash cells away, and (3) an automated micropipette using vacuum-generated fluid flow to lift cells up. These techniques confirmed the results obtained with the high-temporal-resolution Epic BT, but could only provide end-point data. In contrast, complex, nonmonotonic cell adhesion kinetics measured by the high-throughput optical biosensor is expected to open a window on the hidden background of the immune cell–extracellular matrix interactions.

Non-identical twins: Different faces of CR3 and CR4 in myeloid and lymphoid cells of mice and men

“…Studying the spreading of macrophages we found that anti-CR4 also inhibited this function of these human cells. [17,124,125]. Immune cells of the monocytic lineage use specific adhesive structures for cell migration, called podosomes.…”

Single-cell adhesion force kinetics of cell populations from combined label-free optical biosensor and robotic fluidic force microscopy

“…The possibility to chemically modify the sensing surface allows for the examination of specific molecular interactions, such as Arg-Gly-Asp tripeptide (RGD motif) mediated activation and clustering of integrin proteins. The high-quality data provided by this technique have found its use in various fields such as receptor biology [49][50][51] , immune cell biology 52 , environmental toxicology 53 , or the testing of natural active compounds 22,[54][55][56] . Depending on their configuration, RWG sensors are generally compatible with standard 96-or 384-well plates, providing a platform for high-throughput measurements, where numerous cell populations can be measured at the same time under different conditions.…”

Green tea polyphenol tailors cell adhesivity of RGD displaying surfaces: multicomponent models monitored optically

“…The copolymer and its cell adhesive, functionalized counterpart, PLL- g -PEG-RGD can be mixed to vary the cell adhesion ligand density at the nanoscale, and to induce cell adhesion and spreading on the coated surfaces. Here, we use the mixture of PLL- g -PEG and PLL- g -PEG-RGD to control the spreading abilities of the monitored cells 26 27 . Note, PLL- g -PEG is often considered as a gold standard of antifouling surfaces, but the interaction of these coatings with polyphenols has never been investigated.…”