Cells were plated in collagen (0.3 mg/mL)-coated petri dishes for 24 h and placed in a petri dish heater on the AFM stage in 25 mM HEPES-containing media, for a maximum of one hour. Cells incubated with Withaferin-A were measured between 2.75 and 3.25 h of incubation with the drug. The AFM used was a JPK NanoWizard 4. Bruker MSNL-10 cantilevers with a nominal spring constant of 0.03 N/m were used with a setpoint of 3 nN. Measurements were taken in QI mode with a 300 µm/s cantilever speed over a grid of 32 by 32 pixels that covered the whole area of the cell. Data was processed using an in-house MATLAB pipeline which uses a contact point detection method optimised for adherent cells [33 (link),34 (link)]. Since we found a large variability between cell mechanical properties between experiments on different days, results were from a single experiment designed to maximise cell numbers captured in one day.
Msnl 10 cantilevers
Manufactured by Bruker
Sourced in United States
The MSNL-10 cantilevers are high-performance atomic force microscopy (AFM) probes designed for sensitive topography measurements. They feature a stiff silicon nitride cantilever with a sharp, super-durable diamond-like carbon (DLC) tip. The MSNL-10 cantilevers are optimized for non-contact and tapping-mode AFM imaging.
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Lab products found in correlation
6 protocols using msnl 10 cantilevers
1
AFM Measurements of Withaferin-A Effects
2
Tapping Mode AFM Imaging of Proteins
3
Atomic Force Microscopy of Pre-wetted TIPS Microspheres
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Pre-wetted TIPS microspheres were mounted on sterile glass slides using super glue and imaged under deionized water in intermittent contact (liquid) mode employing a JPK NanoWizard II AFM (JPK Instruments, Germany) mounted on an Olympus IX71 inverted microscope. MSNL-10 cantilevers (Bruker, Santa Barbara, USA), tuned to a drive frequency of ∼46 kHz (nominal resonant frequency 25–50 kHz), were employed with a constant line rate of 0.5 Hz. Gain parameters and set point were adjusted according to each sample. Thermal resonance calibration yielded a cantilever spring constant of 0.11 N/m (nominal value 0.1 N/m). After focusing on an area of interest, images with different scan sizes of 5 × 5 µm were obtained at random sites of each sample and processed with the Gwyddion 2.37 SPM software.
4
Biofilm Characterization by AFM Imaging
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A JPK Nanowizard 1 AFM (JPK Instruments Ltd, Germany) was used to obtain example 50 × 50 µm, 10 × 10 µm and 3 × 3 µm AFM images from each biofilm covered HAP disc under PBS conditions. These were conducted using MSNL-10 cantilevers (Bruker Ltd, France) after 1 hour in PBS.
5
Nanoalgosomes Characterization by AFM
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Atomic Force Microscopy images were captured by using a Nanowizard III scanning probe microscope (JPK instruments, AG Germany) equipped with a 12 μm scanner. Nanoalgosomes were initially concentrated by ultracentrifugation and resuspended in MilliQ water to a final concentration of 5 × 1011 particles/ml, as previously estimated by NTA.
For measurements on dry samples, a 30 μl drop of the samples was directly deposited on freshly cleaved mica, incubated for 10 min, and then gently dried under nitrogen flow. Measurements were performed in tapping mode by using a NSC‐15 (Mikromasch) cantilever (spring constant 40 N/m, typical tip radius 8 nm). Measurements with softer cantilevers (data not shown) were carried out with MSNL‐10 cantilevers (Bruker; lever D, spring constant 0.03 N/m, nominal tip radius 2 nm).
For measurements on dry samples, a 30 μl drop of the samples was directly deposited on freshly cleaved mica, incubated for 10 min, and then gently dried under nitrogen flow. Measurements were performed in tapping mode by using a NSC‐15 (Mikromasch) cantilever (spring constant 40 N/m, typical tip radius 8 nm). Measurements with softer cantilevers (data not shown) were carried out with MSNL‐10 cantilevers (Bruker; lever D, spring constant 0.03 N/m, nominal tip radius 2 nm).
6
AFM Analysis of Candida albicans Adhesion
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All AFM imaging and nanomechanic experiments were carried out with a JPK Nanowizard system. Images were obtained with intermittent contact mode in fluid; by employing MSNL-10 cantilevers (Bruker, USA) with a spring constant of 0.1 N/m. Gain parameters were adjusted during imaging to allow for optimum image acquisition. Images of C. albicans cells and hyphae were acquired at different magnifications with 256x256 pixels and a scan rate of 0.3 Hz. SCFS experiments were carried out with a loading force of 1 nN, a constant speed of 2 µm/s, and surface delay times of 0, 1, 5, 10 and 30 s. For yeast cells and the budding cells, force curves were obtained on the centre of the cell as observed with the optical microscope (Figure 1). For hyphal tubes, force curves were performed in the middle portion of the tube (Figure 1). All SCFS experiments were carried out in PBS buffer, maintaining probes and C. albicans-coated surfaces fully hydrated throughout the whole process. Force curves obtained over a total of 7 independent yeast cells and hyphae were utilised for data analysis.
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