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Dissociation Constant of Integrin-RGD Binding in Live Cells from Automated Micropipette and Label-Free Optical Data

Authors :
Tamás Gerecsei
Péter Chrenkó
Nicolett Kanyo
Beatrix Péter
Attila Bonyár
Inna Székács
Balint Szabo
Robert Horvath
Source :
Biosensors, Vol 11, Iss 2, p 32 (2021)
Publication Year :
2021
Publisher :
MDPI AG, 2021.

Abstract

The binding of integrin proteins to peptide sequences such as arginine-glycine-aspartic acid (RGD) is a crucial step in the adhesion process of mammalian cells. While these bonds can be examined between purified proteins and their ligands, live-cell assays are better suited to gain biologically relevant information. Here we apply a computer-controlled micropipette (CCMP) to measure the dissociation constant (Kd) of integrin-RGD-binding. Surface coatings with varying RGD densities were prepared, and the detachment of single cells from these surfaces was measured by applying a local flow inducing hydrodynamic lifting force on the targeted cells in discrete steps. The average behavior of the populations was then fit according to the chemical law of mass action. To verify the resulting value of Kd2d = (4503 ± 1673) 1/µm2, a resonant waveguide grating based biosensor was used, characterizing and fitting the adhesion kinetics of the cell populations. Both methods yielded a Kd within the same range. Furthermore, an analysis of subpopulations was presented, confirming the ability of CCMP to characterize cell adhesion both on single cell and whole population levels. The introduced methodologies offer convenient and automated routes to quantify the adhesivity of living cells before their further processing.

Details

Language :
English
ISSN :
11020032 and 20796374
Volume :
11
Issue :
2
Database :
Directory of Open Access Journals
Journal :
Biosensors
Publication Type :
Academic Journal
Accession number :
edsdoj.f7ee72160eb6400b9010d4216082e7d3
Document Type :
article
Full Text :
https://doi.org/10.3390/bios11020032