AFM analysis was performed as described previously [82 (link)]. Shortly, amyloid samples were prepared by applying a drop of 10 μL medium on freshly cleaved mica, V1 grade (NanoAndMore GmbH, Germany). After incubation for 10 min, the sample was rinsed with deionised water (Merck Millipore Inc., Burlington, MA, USA) and dried under a gentle stream of argon. A Multimode 8 Nanoscope atomic force microscope (AFM, Bruker, Billerica, MA, USA) was used to image the surfaces of the mica substrate and the deposited amyloid structures.
Multimode 8 nanoscope atomic force microscope
Manufactured by Bruker
Sourced in United States
The Multimode 8 Nanoscope atomic force microscope is a high-resolution imaging system designed for surface characterization at the nanoscale. It utilizes a cantilever-based probe to scan the sample surface, providing detailed topographical information and surface property data.
Automatically generated - may contain errors
5 protocols using multimode 8 nanoscope atomic force microscope
1
Atomic Force Microscopy of Amyloid
2
Atomic Force Microscopy Imaging of Samples
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A Multimode 8 Nanoscope atomic force microscope (AFM, Bruker, Billerica, MA, U.S.A.) was used to image the samples’ surfaces. Silicon cantilevers with a spring constant of ca. 5 Nm−1 (TapDLC-150, BudgetSensors, Sofia, Bulgaria) were applied for imaging in PeakForce TappingTM microscopy mode. The sample preparation procedure includes the application of 150 µL of solution on the clean mica plate, V1 grade (NanoAndMore GmbH, Wetzlar, Germany). After 10 min, the plates were rinsed with deionized water and dried under a gentle stream of argon.
3
Imaging Oligomeric Structures on Mica by AFM
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Multimode 8 Nanoscope atomic force microscope (AFM, Bruker, USA) was used to image the surfaces of the mica substrate and the freshly deposited oligomeric structures. Silicon cantilevers, ETALON series with a spring constant of ca. 3.5 Nm−1 (NT-MDT, Russia) or HQ:NSC19/No Al type with a spring constant of ca. 0.5 Nm−1 (Mikromasch, Bulgaria) were applied for imaging in PeakForce Tapping™ Force Microscopy Mode. Calibration of the microscope was achieved by the imaging of calibration gratings supplied by the manufacturer. The images presented in this work are height type images. The examination of surfaces for artefacts by AFM, and the reproducibility, was performed in the common way, i.e. by changing the AFM cantilever and moving the sample in the X or Y direction or by varying the scanning angle and scan rate. Oligomeric samples were prepared by applying a drop of 10 μl Aβ1–42 solution on freshly cleaved mica (Ted Pella Inc., USA). After incubation for 10 min, the sample was rinsed with deionised water (Millipore Inc., USA) and dried under a gentle stream of argon.
4
Imaging Crosslinked Nucleosome Arrays
5
Atomic Force Microscopy of Crosslinked Tetranucleosomes
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Purified tetranucleosomes were diluted to 10 nM and crosslinked with 1% formaldehyde for 1 h at room temperature. Crosslinked sample was dialyzed against 20 mM HEPES-NaOH pH 7.5; 1 mM EDTA; 1 mM DTT, and centrifuged at 20,000 x g to remove aggregates. Crosslinked tetranucleosome sample was diluted to 1 nM using 10 mM MOPS pH 7.0 and 5 mM MgCl2, and 3 µL of sample were deposited and incubated for 2 min on freshly cleaved bare mica V1 (Ted Pella Inc.), after which was rinsed with Milli-Q water, and then gently dried under a stream of N2 perpendicular to the mica surface. AFM micrographs were taken with a MultiMode NanoScope 8 atomic force microscope (Bruker Co.) equipped with a vertical engagement scanner E. The samples were excited at their resonance frequency (280-350 kHz) with free amplitudes (Ao) of 2-10 nm and imaged in tapping mode using silicon cantilevers (Nanosensors). The image amplitude (set point As) and A0 ratio (As/A0) was kept at ~0.8 in a repulsive tip-sample interaction regime, and phase oscillations were no greater than ± 5 degrees. The surface was rastered following the fast scan axis (x) at rates of 2 Hz, capturing the retrace line to reconstruct the AFM micrographs. All samples were scanned at room temperature in air, at a relative humidity of 30%.
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