Measuring the adhesion limit of fibronectin for fibroblasts with a narrow-gap rotational rheometer

Abstract

We study the adhesion limit of 3T6 fibroblasts, cultured in Dulbecco's modified Eagle's medium with 10% fetal bovine serum at 37 °C and 5% CO, with a narrow-gap rotational rheometer in the parallel-disk configuration. Reducing the uncertainty in gap width to about 1 µm allows studying the cells at narrow gaps, which enables to study the critical shear stress of the cells in low-viscous media. The adhesion limit on fibronectin-coated glass plates is determined as a function of concentration and adhesion time. We found that cells in groups have a tendency to detach at slightly higher shear stresses than single cells. Moreover, 60 min after the settling phase are enough for the cells to adhere to the coated plate at maximum strength. We show that the setup may also be used for cells that are not adhered from suspension, but are grown directly on the substrate.

Quantifying cell adhesion through forces generated by acoustic streaming

“…5 . The trend is consistent with previous research [35] . Further, experiments were performed to evaluate the cell adhesion with different culture and PBS immersion times.…”

Narrow-Gap Rheometry: A Novel Method for Measuring Cell Mechanics

“…This is far below the reported effective zero-gap error of commercial rheometers of 25 µm to 70 µm [86], which, apart from being due to disk roughness and residual disk inclination, can be caused by squeeze flow during zeroing the rheometer [85]. The enhanced precision allows the extending of the measurement window to much higher shear rates [87,88] as well as studying the adhesion [89] and load limits [90] of cells in low viscous environments. With their device, Dakhil et al reached the linear viscoelastic range at small deformations, where the stress amplitude is proportional to the deformation amplitude [20,91].…”

Paper‐Based Cell Cryopreservation

“…As such, short attachment times (<60 minutes) result in formation of fewer cell-to-surface bonds because of the round shape of the cells. Hence, when force is applied these bonds are easily dissociated resulting in detachment of cells 35,37 . Taken together, we hypothesize that in our method fibronectin acts more like a lubricant to enhance the cell release from the paper, which is associated with their short-length (<1 minute) cell-substrate bond strengths 29,31,37,38 .…”

“…Hence, when force is applied these bonds are easily dissociated resulting in detachment of cells 35,37 . Taken together, we hypothesize that in our method fibronectin acts more like a lubricant to enhance the cell release from the paper, which is associated with their short-length (<1 minute) cell-substrate bond strengths 29,31,37,38 . In addition, we assume that the highly (>50 µm) 3D porous structure of the paper 22,39 also enhances the release of cells upon shear.…”