Omcl tr400pb 1

The electrical properties of EB-P3HT and a chemically synthesized regioregular P3HT reference were measured with current sensing-atomic force microscopy (CS-AFM, 5500AFM; Agilent Technologies). The P3HT sample (10 × 15 × 2 µm) was prepared on a glass substrate coated with a vapor deposited Al film. Au-coated cantilevers (OMCL-TR400PB-1, Olympus Corporation; spring constant: 0.09 N m⁻¹) were used to measure the current between the tip and the Al film. The EB-P3HT was doped with iodine by immersion in an aqueous solution containing 0.1 M KI and I₂ for 1 h.

To visualize the peptide bactericidal effects on bacterial cells, AFM was performed. In AFM experiments, the bacterial cells were washed twice with PBS and treated with peptides. After the peptide treatments, 10 mL of the bacterial suspension was placed onto a freshly cleaved mica disc (Ted Pella, Redding, CA, USA), and samples were allowed to be adsorbed for 20–40 min. The mica discs were then gently rinsed twice with Milli Q water and dried in a desiccator. The prepared samples were imaged using an AFM instrument (Nanowizard, JPK Instruments, Germany) operating in contact mode. Images were collected at a resolution of 512 × 512 pixels using a scanning speed of 1.0–2.0 Hz. The AFM probes used were oxide-sharpened silicon nitride (Si₃N₄) probes (OMCL-TR400PB-1, Olympus, Japan) with a spring constant of 0.02 N/m. The SPM image processing package version 3.1.6 (JPK Instruments) was used for AFM image data processing and analysis.

A two-terminal method was used to obtain the contact resistance at the CNT/Au interface, so the robotic system was configured with two manipulators based on a FE-SEM apparatus (JSM-6500F, JEOL), as shown schematically in Figure 1. Both of the manipulators were driven by picomotors (8301-UHV, Newport) in x-y-z directions with a resolution of 30 nm. Two AFM cantilevers covered with layers of Au (OMCL-TR400PB-1, OLYMPUS) were mounted on the manipulators as the end effectors. A MWCNT forest prepared by Arc charge method was placed on top of the sample stage. Tungsten hexacarbonyl (W(CO)₆, MKBR3026V, SIGMA-ALDRICH) was introduced into the specimen chamber as the precursor for EBID with tungsten. A visual-based force feedback system was developed with the robotic system for real-time manipulation (not shown in Figure 1).
The AFM cantilevers were able to be mounted in different orientations to achieve different tasks. Horizontally fixing the two AFM cantilevers allowed the total resistance to be measured, as shown in Figure 2. Changing the orientation of one cantilever to vertical orientation allowed the contact force to be characterized, as shown in Figure 3(a).

All images were collected using an AFM (Nanowizard^TM, JPK instruments, Berlin, Germany) installed on an inverted optical microscope (Nikon Corporation, Tokyo, Japan). The AFM probes used were oxidize-sharpened silicon nitride probes (OMCL-TR400PB-1, Olympus, Tokyo, Japan) with a spring constant of 0.02 N/m. Ten microliters of samples treated with or without gramicidin A or Ca²⁺ ions at the designed concentration were placed on a cleaved mica disc (Ted Pella Inc., Redding, CA, USA) for 20–40 min. The mica disc was then rinsed twice with distilled water and dried for 24 h. The dried samples were imaged in an AFM operating in a contact mode. Images were acquired at a scanning rate of 1–2 Hz and a resolution of 512 × 512 pixels. Image processing and analysis were performed using SPM software v. 3.16 (Nanowizard^TM, JPK instruments, Berlin, Germany, Germany).

A certain magnitude of mechanical force was applied to the fixed myotubes using a flexible glass needle as described previously^{20 (link)}. The needle was made from a glass rod using a glass electrode puller (P-1000; Sutter Instrument, CA, U.S.A). The tip of the needle was made to be approximately 5 µm in diameter. The bending stiffness of the needle was determined to be 465 nN/µm; this was measured from the displacement of the tip of the needle when a constant force was applied to the needle using an atomic force microscopy cantilever with a precisely calibrated stiffness (OMCL-TR400PB-1; Olympus, Tokyo, Japan). Myotubes on the 5th day of differentiation were fixed with 1% glutaraldehyde in PBS for 10 min at room temperature and washed with 30 mmol/L glycine in PBS. The needle was pricked into a fixed myotube using a micromanipulator (MWO-3; Narishige, Tokyo, Japan) to apply mechanical force, and observed under the phase-contrast microscope. The deflection of the needle was measured as the distance between the tips of the needles during and after the application of force. The force applied to the myotubes was calculated as the product of the bending stiffness (nN/µm) and the deflection (µm) of the needle. All data were subjected to outlier tests and the data excluding outliers were plotted graphically.