On-substrate lysis of cells was performed based on a previous study by Kim et al. [21 (link)] with modifications. S. solfataricus culture were harvested, washed, and deposited on a clean coverslip for 10 min in room temperature and dried using nitrogen gas. Cells were lysed through the addition of lysis buffer (10 mM Tris-Cl, pH 7.5, 10 mM EDTA, 0.5% Triton-X), followed by washing with sterile distilled/deionized water and drying with nitrogen gas. Samples were then imaged through atomic force microscopy using the XE-BIO system (Park Systems, Suwon, South Korea).
Xe bio system
Manufactured by Park Systems
Sourced in Cameroon
The XE-Bio system is a versatile lab equipment designed for advanced biological and molecular analysis. It provides high-resolution imaging and measurement capabilities for a wide range of applications.
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7 protocols using xe bio system
1
On-substrate Lysis and AFM Imaging
2
Non-invasive Myotube Surface Imaging
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Surface structures of the differentiated myotubes cultured on the PEDOT/MWCNT sheet were obtained using the SICM nano-imaging technique43 . SICM images were obtained with the XE-Bio System (Park Systems). Myotube samples were embedded in phosphate-buffered saline (PBS) solution and placed on an x-y flat scanner stage (XE-Bio System) mounted on an inverted microscope (IX71, Olympus). The SICM probe glass nano-pipette (inner diameter of ~100 nm) was fabricated from borosilicate glass tubing (Warner Instruments) using a CO2-laser-based micropipette puller (P-2000, Sutter Instruments, CA) and filled with PBS electrolyte solution. Using the approach-retract scanning (ARS) or hopping modes of SICM imaging43 , 3D cell surface images of the live myotubes were successfully obtained in a non-invasive manner.
3
Nanoscale Surface Charge Density Mapping
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Surface charge density (SCD) measurements were performed using a scanning ion conductance microscope (SICM) setup on an XE-Bio system (Park Systems, Suwon, South Korea) using Ag/AgCl electrodes. The imaging buffer consisted of 10 mM HEPES (pH 7.0) and 150 mM KCl. A single borosilicate glass nanopipette was used for all measurements. SCD sensitive current-distance curves were obtained at an approach and withdraw speed of 200 nm s−1 at potentials of ±0.5 V. For each sample, current curves obtained at ±0.5 V were first normalized with respect to the bulk current. Following this, the two curves were subtracted from each other, (+)100 mV-(−)100 mV yielding ΔI_N. Multiple curves were obtained at various locations for verifying the curve tendencies. The curves shown are representative of the SCD properties.
4
Atomic Force Microscopy of Aortic Valve
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An XE-Bio system (Park systems, Santa Clara, CA, USA) in contact mode was used for all AFM measurements. Pre-calibrated gold-coated bead AFM probe (Novascan Technologies, Ames IA, USA) with a 1 μm diameter SiO2 microsphere were used for the indentation measurements. The spring constant of the tip was 0.1 N/m. Force curves measurements at the targeted region of AoV leaflet were performed with a probe approaching speed of 0.3 μm/s and peak force range 0.4–2.4nN. Each targeted point was measured three to five times to ensure the accuracy of the measurement.
5
Characterizing NP Morphology via SEM and AFM
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The morphology of NPs was characterized by scanning electron microscopy (SEM, JEOL JSM-7800F, Akishima, Tokyo, Japan) and atomic force microscopy (AFM, XE-Bio system, Park Systems Corp, Su-won, Korea). SEM images were obtained with a magnification of 30,000× using an acceleration voltage of 5.0 kV. AFM images were reconstructed in the non-contact mode using Nanosensors: NCHR cantilevers (force constant 10 to 130 N/m). The analysis was performed using 5 × 5 μm scanning images. The three-dimensional (3D) images were analyzed with the software Gwyddion version 2.49 available online at http://gwyddion.net/ .
6
SICM Imaging of DNA Samples
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After imaging of DNA using AFM, the sample was moved to an XE‐Bio system (Park Systems, Suwon, South Korea) setup to record SICM images. An imaging buffer consisting of Tris (10 mM), acetic acid, pH 8.0 and KCl (150 mM) was used throughout all experiments. Imaging was conducted at +100 mV (with respect to the bath electrode) with a set‐point of 99 % of the free current (338 pA).
7
SICM Imaging with XE-Bio System
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All SICM images were recorded using an XE-Bio system (Park Systems, Suwon, South Korea). The instrumentation has been described previously23 (link). The setup uses a Femto DLPCA 200 amplifier and a Nikon Eclipse TI-U optical microscope.
The imaging buffer consisted of 150 mM NaCl (Sigma, USA) and 10mM HEPES (Sigma, USA) in milli-Q grade water adjusted to pH 7. During imaging the temperature was kept at 20 °C. The SICM was operated in constant current mode (DC). Before each scan the unperturbed current far from the surface was measured and a 1%-drop setpoint calculated. This low setpoint (99%) was chosen to optimize the signal to noise ratio, while higher setpoints (99.5–99.7%) are often used for live-cell imaging to prevent tip–sample collisions41 (link). After the measurement the unperturbed current was measured again. Scans with a drift in unperturbed current of more than 0.3% were discarded. Images were scanned subsequently with bias potentials of +100 and −100 mV. A period of 2–5 min was introduced following a change of the potential to assure a stable current.
The imaging buffer consisted of 150 mM NaCl (Sigma, USA) and 10mM HEPES (Sigma, USA) in milli-Q grade water adjusted to pH 7. During imaging the temperature was kept at 20 °C. The SICM was operated in constant current mode (DC). Before each scan the unperturbed current far from the surface was measured and a 1%-drop setpoint calculated. This low setpoint (99%) was chosen to optimize the signal to noise ratio, while higher setpoints (99.5–99.7%) are often used for live-cell imaging to prevent tip–sample collisions41 (link). After the measurement the unperturbed current was measured again. Scans with a drift in unperturbed current of more than 0.3% were discarded. Images were scanned subsequently with bias potentials of +100 and −100 mV. A period of 2–5 min was introduced following a change of the potential to assure a stable current.
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