title: Measurement of cell adhesion force by vertical forcible detachment using an arrowhead nanoneedle and atomic force microscopy.
abstract: The properties of substrates and extracellular matrices (ECM) are important factors governing the functions and fates of mammalian adherent cells. For example, substrate stiffness often affects cell differentiation. At focal adhesions, clustered-integrin bindings link cells mechanically to the ECM. In order to quantitate the affinity between cell and substrate, the cell adhesion force must be measured for single cells. In this study, forcible detachment of a single cell in the vertical direction using AFM was carried out, allowing breakage of the integrin-substrate bindings. An AFM tip was fabricated into an arrowhead shape to detach the cell from the substrate. Peak force observed in the recorded force curve during probe retraction was defined as the adhesion force, and was analyzed for various types of cells. Some of the cell types adhered so strongly that they could not be picked up because of plasma membrane breakage by the arrowhead probe. To address this problem, a technique to reinforce the cellular membrane with layer-by-layer nanofilms composed of fibronectin and gelatin helped to improve insertion efficiency and to prevent cell membrane rupture during the detachment process, allowing successful detachment of the cells. This method for detaching cells, involving cellular membrane reinforcement, may be beneficial for evaluating true cell adhesion forces in various cell types.
authors: Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588, Japan.
authors: Seunghwan
authors: S
authors: Ryu
authors: Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588, Japan.
authors: Yui
authors: Y
authors: Hashizume
authors: Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588, Japan.
authors: Mari
authors: M
authors: Mishima
authors: Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology, Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan.
authors: Ryuzo
authors: R
authors: Kawamura
authors: Faculty of Medicine, University of Tsukuba, 1-1-1 Ten-nohdai, Tsukuba, Ibaraki 305-8573, Japan.
authors: Masato
authors: M
authors: Tamura
authors: Faculty of Medicine, University of Tsukuba, 1-1-1 Ten-nohdai, Tsukuba, Ibaraki 305-8573, Japan.
authors: Hirofumi
authors: H
authors: Matsui
authors: Department of Applied Chemistry, Graduate School of Engineering, Osaka University, 2-1 Yamada-oka, Suita, Osaka 565-0871, Japan.
authors: Michiya
authors: M
authors: Matsusaki
authors: Department of Applied Chemistry, Graduate School of Engineering, Osaka University, 2-1 Yamada-oka, Suita, Osaka 565-0871, Japan.
authors: Mitsuru
authors: M
authors: Akashi
authors: Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588, Japan; Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology, Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan. Electronic address: chikashi-nakamura@aist.go.jp.
authors: Chikashi
authors: C
authors: Nakamura
mesh_terms: D000818:Animals; D001703:Biophysics / Q000295:instrumentation / Q000379:methods*; D002448:Cell Adhesion; D002460:Cell Line; D002462:Cell Membrane; D051379:Mice; D018625:Microscopy, Atomic Force / Q000295:instrumentation / Q000379:methods*; D036103:Nanotechnology / Q000295:instrumentation*; D051381:Rats
keywords: AFM; Arrowhead nanoneedle; Cell adhesion force; Nanofilm