title: Stability enhancement of an atomic force microscope for long-term force measurement including cantilever modification for whole cell deformation.
abstract: Atomic force microscopy (AFM) is widely used in the study of both morphology and mechanical properties of living cells under physiologically relevant conditions. However, quantitative experiments on timescales of minutes to hours are generally limited by thermal drift in the instrument, particularly in the vertical (z) direction. In addition, we demonstrate the necessity to remove all air-liquid interfaces within the system for measurements in liquid environments, which may otherwise result in perturbations in the measured deflection. These effects severely limit the use of AFM as a practical tool for the study of long-term cell behavior, where precise knowledge of the tip-sample distance is a crucial requirement. Here we present a readily implementable, cost effective method of minimizing z-drift and liquid instabilities by utilizing active temperature control combined with a customized fluid cell system. Long-term whole cell mechanical measurements were performed using this stabilized AFM by attaching a large sphere to a cantilever in order to approximate a parallel plate system. An extensive examination of the effects of sphere attachment on AFM data is presented. Profiling of cantilever bending during substrate indentation revealed that the optical lever assumption of free ended cantilevering is inappropriate when sphere constraining occurs, which applies an additional torque to the cantilevers "free" end. Here we present the steps required to accurately determine force-indentation measurements for such a scenario. Combining these readily implementable modifications, we demonstrate the ability to investigate long-term whole cell mechanics by performing strain controlled cyclic deformation of single osteoblasts.
authors.affiliation: Department of Mechanical and Biomedical Engineering, National University of Ireland, Galway, Ireland.
authors.forename: P P
authors.initials: PP
authors.lastname: Weafer
authors.forename: J P
authors.initials: JP
authors.lastname: McGarry
authors.forename: M H
authors.initials: MH
authors.lastname: van Es
authors.forename: J I
authors.initials: JI
authors.lastname: Kilpatrick
authors.forename: W
authors.initials: W
authors.lastname: Ronan
authors.forename: D R
authors.initials: DR
authors.lastname: Nolan
authors.forename: S P
authors.initials: SP
authors.lastname: Jarvis
mesh_terms: D000818:Animals; D001696:Biomechanical Phenomena; D002138:Calibration; D002470:Cell Survival; D055595:Mechanical Phenomena; D018625:Microscopy, Atomic Force / Q000295:instrumentation*; D013997:Time Factors