Successive detection of insulin-like growth factor-II bound to receptors on a living cell surface using an AFM.
In this study, we have developed a method of mechanical force detection for ligands bound to receptors on a cell surface, both of which are involved in a signal transduction pathway. This pathway is an autocrine pathway, involving the production of insulin-like growth factor-II (IGF-II) and activation of the IGF-I receptor, involved in myoblast differentiation induced by MyoD in C3H10T1/2 mouse mesenchymal stem cells. Differentiation of C3H10T1/2 was induced with the DNA demethylation agent 5-azacytidine (5-aza). The etched AFM tip used in the force detection had a flat surface of which about 10 µm(2) was in contact with a cell surface. The forces required to rupture the interactions of IGF-IIs on a cell and anti mouse IGF-II polyclonal antibody immobilized on an etched AFM tip were measured within 5 days of induction of differentiation. The mean unbinding force for a single paired antibody-ligand on a cell was about 81 pN, which was measured at a force loading rate of about 440 nN/s. The percentage of unbinding forces over 100 pN increased to 32% after 2 days from the addition of 5-aza to the medium. This method could be used in non-invasive and successive evaluation of a living cell's behavior.
Han
Sung-Woong
SW
Research Institute for Cell Engineering (RICE), National Institute of Advanced Industrial Science and Technology (AIST), Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan.
Mieda
Shingo
S
Nakamura
Chikashi
C
Kihara
Takanori
T
Nakamura
Noriyuki
N
Miyake
Jun
J
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001374:Azacitidine / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D003584:Cytological Techniques; D007335:Insulin-Like Growth Factor II / Q000378:metabolism*; D051379:Mice; D018625:Microscopy, Atomic Force; D017526:Receptor, IGF Type 1 / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism
D000911:Antibodies, Monoclonal; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D007335:Insulin-Like Growth Factor II; D017526:Receptor, IGF Type 1; D001374:Azacitidine