Abstract
Integrin activation is a crucial event for multiple biological functions. Therefore, methods to detect integrin activation are vital. Since the main cellular function of integrins is adhesion, we and others utilize this feature to measure integrin activation. Here, we describe how to detect the activity of the fibronectin-binding integrin α5β1 using a fusion of glutathione S-transferase (GST) to the 9th, 10th, and 11th type III repeats on fibronectin (GST-FNIII9-11). Moreover, we detail how to measure αvβ3 integrin activity using the ligand-mimetic WOW-1 antibody that selectively binds unoccupied αvβ3 integrins. Finally, we describe methods of testing ligation of fibronectin-binding integrins utilizing monoclonal antibodies against ligand-induced binding sites (LIBS).
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Acknowledgments
This work was supported by National Heart, Lung, and Blood Institute R01 HL098435, HL133497, HL141155, and GM121307 (to A.W.O.), by an American Heart Association Predoctoral Fellowship (19PRE34380751) and Malcolm Feist Cardiovascular Research Endowment Predoctoral Fellowship (to Z.A.Y.).
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Al-Yafeai, Z., Orr, A.W. (2021). Quantification of Integrin Activation and Ligation in Adherent Cells. In: Vicente-Manzanares, M. (eds) The Integrin Interactome. Methods in Molecular Biology, vol 2217. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0962-0_2
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DOI: https://doi.org/10.1007/978-1-0716-0962-0_2
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