Automated AFM force curve analysis for determining elastic modulus of biomaterials and biological samples

Abstract

The analysis of atomic force microscopy (AFM) force data requires the selection of a contact point (CP) and is often time consuming and subjective due to influence from intermolecular forces and low signal-to-noise ratios (SNR). In this report, we present an automated algorithm for the selection of CPs in AFM force data and the evaluation of elastic moduli. We propose that the CP may be algorithmically easier to detect by identifying a linear elastic indentation region of data (high SNR) rather than the contact point itself (low SNR). Utilizing Hertzian mechanics, the data are fitted for the CP. We first detail the algorithm and then evaluate it on sample polymeric and biological materials. As a demonstration of automation, 64 x 64 force maps were analyzed to yield spatially varying topographical and mechanical information of cells. Finally, we compared manually selected CPs to automatically identified CPs and demonstrated that our automated approach is both accurate (< 10 nm difference between manual and automatic) and precise for non-interacting polymeric materials. Our data show the algorithm is useful for analysis of both biomaterials and biological samples.

Tissue and cellular biomechanics during corneal wound injury and repair

“…Tissue samples were mounted for AFM without any glue and analyses performed as described previously [33–36]. Briefly, elastic modulus was determined in contact mode using a silicon nitride probe with a square pyramid tip incorporated at the free end ( κ =0.32 N/m; PNT-TR-50, Nano and More, Lady’s Island, SC); a 5 µm radius borosilicate bead was attached to the pyramid of the cantilever and contact-mode mechanics was determined by obtaining five force-versus-indentation curves at five difference locations of the tissue.…”

“…Cells were cultured on the hydrogels, in DMEM low glucose supplemented with 10% FBS and 1% P/S/F, for 3 days in the presence or absence of 10 ng/ml TGFβ1 for rabbit stromal cells or 2 ng/ml TGFβ1 for human stromal cells [31]. Cell mechanics of stromal cells (at least 10 cells per condition, with 5 force curves per cell) cultured on the matrices of differing stiffness were determined by AFM as previously described [36, 39]. Experiments were performed in triplicate.…”

Investigation of the AFM contact-mode force calibration with simulation

“…The Young's modulus can be obtained from the proportional factor by fitting the selected theoretical model on the measured d(z) curve. In practice the most common way to determine the V/nm conversion factor (also called the deflection inverse optical lever sensitivity, or DeflInvOLS [8]) is performing contact-mode point-spectroscopy measurements on an ideally hard substrate. The theoretical background is, that during pointspectroscopy, in the case of a hard material, the deflection of the cantilever in nm (d) ideally equals the movement of the tip in the Z dimension in nm (z), thus the gradient of the obtained point-spectroscopy curve is the inverse of the conversion factor (Fig.…”

Searching events in AFM force‐extension curves: A wavelet approach

“…In particular, in the AFM F-z curves analysis, several approaches have been made to implement algorithms for the automatic extraction of features from the measurements (Andreopoulos & Labudde, 2011;Benítez et al, 2013;Chang et al, 2014;Lin, Dimitriadis, & Horkay, 2007a,b;Scholl & Marszalek, 2014).…”