All Raman spectra were processed by removing signals attributed to cosmic rays and subtracting the baseline (Wire 5.2 software, Renishaw, UK). A component analysis (with normalisation by subtraction of minimum value and scaling to unit variance; Wire 5.2 software, Renishaw, UK) was applied to each sample spectrum to create maps of mineral distribution and estimate mineral abundance in the sample. The component analysis used reference spectra of ferrihydrite, lepidocrocite, goethite and hematite (hematite produced by beam damage and used to check for beam damage, details in Section 9 of the ESI† ). The size and distribution of regions with distinct mineral identity in the 1 μm-resolution μ-Raman spectral component analysis maps were measured using image processing techniques. Binary images were produced by identifying pixels with mineral content above 50% of the normalised total range measured in the sample. Image segmentation analysis on the binary images employed an unsupervised watershed model with 3 μm minimum distance between particle centres, implemented with the SciPy and Scikit-image analysis packages.84,85 (link) Particle dimensions were measured with Scikit-image85 (link) and nearest neighbour distances were calculated using a ball-tree model from the Scikit-learn package.86
Wire 5
Manufactured by Renishaw
Sourced in United Kingdom
Wire 5.2 software is a tool used for the control and monitoring of Renishaw's wire EDM (Electrical Discharge Machining) machines. It provides an interface for operators to configure and manage the machining process.
Automatically generated - may contain errors
Lab products found in correlation
Invia confocal raman microscope, by Renishaw (4 mentions)
Invia qontor, by Renishaw (2 mentions)
Invia reflex, by Renishaw (2 mentions)
Invia raman spectrometer, by Renishaw (1 mentions)
Dpssl laser, by Renishaw (1 mentions)
Invia reflex confocal raman microscope, by Renishaw (1 mentions)
Opus 8, by Bruker (1 mentions)
Invia qontor microscope, by Renishaw (1 mentions)
532 nm laser, by Zeiss (1 mentions)
Objective lens, by Zeiss (1 mentions)
Invia raman microscope, by Renishaw (1 mentions)
15 protocols using wire 5
1
Raman Spectroscopy for Mineral Characterization
2
Raman Spectroscopy Analysis of AmB-Ag in GUVs and C. albicans
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Raman spectroscopy was carried out using an inVia confocal Raman microscope (Renishaw, Kingswood, UK) with an argon laser (Stellar-REN, Modu-Laser™, Centerville, UT, USA) operating at 457 nm (set at 47 μW power at the sample), equipped with a 60× water-immersed objective (Olympus Plan Apo NA = 1.2). Optical images of AmB-Ag containing GUV and C. albicans fungal cells were obtained and analyzed with WiRE 5.5 software (Renishaw, Kingswood, UK). Based on such images, areas of approximately 10 μm × 10 μm for Raman scanning were selected and mapped with 1 μm spatial resolution. For this study, all the images were recorded with light intensity as low as possible. Raman images were acquired using the Renishaw WiRE 5.5 system in high-resolution mapping mode (HR maps). At each point of the Raman image map, the spectra were recorded with about 1 cm−1 spectral resolution (2400 lines/mm grating) in Raman shift spectral region 1000–2500 cm−1 using an EMCCD detection camera Newton 970 from Andor, Belfast, UK. The acquisition time for a single spectrum was 0.5 s for GUV and 1.0 s for C. albicans. All spectra were pre-processed by cosmic ray removal, noise filtering, and baseline correction using WiRE 5.5 software from Renishaw, Kingswood, UK.
3
Micro-Raman Spectroscopy of Bone Implant Interface
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Micro-Raman spectroscopy was performed using a confocal Raman microscope (Renishaw inVia™ Qontor®) equipped with a 633 nm laser and LiveTrack™ focus-tracking technology. The laser was focused down onto the sample surface using a × 100 (0.9 NA) objective. The Raman scattered light was collected using a Peltier-cooled CCD deep depletion NIR enhanced detector behind 1,800 g mm−1. Two regions of interest were defined: (i) within thread (mineralized bone within the first endosteal thread, i.e., below the level of the original cortical bone, to ensure that only de novo-formed bone was analysed) and (ii) native bone (original cortical bone ≥1 mm from the implant surface). From each region of interest, 6 to 8 spectra were collected (8 s integration time and 10 accumulations per spectrum). Background subtraction and cosmic ray removal were performed in Renishaw WiRE 5.2 software (10 (link)).
4
Raman Spectroscopy of Dry Samples
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Raman scattering spectra of dry samples were acquired using Renishaw Invia Raman spectrometer with 785 nm laser confocal microscope at 200–3200 cm−1 wavelength, 1200 L/mm grating, 10 s exposure time, and 1–5% laser power. Baseline correction, minor smoothening (if required) and normalisation of the spectra were conducted using Renishaw WiRE™ 5.2 software.
5
Raman Spectroscopy for Cell Culture and CEX
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All the spectral measurements were performed using the InVia confocal Raman microscope (Renishaw, Wotton-under-Edge, UK) equipped with a 785 nm laser. Prior to spectral measurements, the acquisition settings were optimised in respect to the focal point, sample volume, laser output, and duration of measurements. Measurements of the cell culture were performed in the range of 381 to 1534 cm -1 using a 10% laser power (30 mW), 30 s acquisition time, 5 accumulations, and line-focus using a 5X objective (Leica Microsystems, Wetzlar, Germany). For each sample, 350 µL of culture was spun down to remove cells, and 300 µL of supernatant was used for the acquisition. The data acquisition was performed using polypropylene (PP) 96-well plates (Greiner Bio-one, Stonehouse, UK) using the Microplate mapping option of the WiRe 5.2 software (Renishaw, Wotton-under-Edge, UK). Data acquisition for the CEX samples was performed similarly to the cell culture samples, with a difference in acquisition range (605-1741 cm -1 ). Additional experiments were performed comparing the signal from the PP 96-well plates with the signal from stainless steel plates, which were custom-made. The acquisition was further optimised using a long-distance 50X objective (Leica Microsystems, Wetzlar, Germany), increasing the laser power to 100%, and decreasing the acquisition time to 10 s and 3 accumulations.
6
Raman Spectroscopy of Hydrogel Samples
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The hydrogel samples were measured using an inVia™ confocal Raman microscope from Renishaw (Gloucestershire, UK) with a 633 nm helium-neon laser. The laser power was controlled at 7.1 mW, and each measurement was conducted with three accumulations. Raman spectra were evaluated using WiRE 5.0 software by Renishaw (Gloucestershire, UK). All spectra were filtered using a Savitzky–Golay polynomial with an order of three and frame length of 31, background subtracted with a polynomial with an order of five, and normalized via total scanning areas from 100 cm−1 to 3600 cm−1, followed by smoothing using a moving mean with a window of 17.
7
Raman Spectroscopic Analysis of Gelatin Samples
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Raman measurements were carried out using the inVia™ confocal Raman microscope (Renishaw, Wotton-under-Edge, UK) equipped with a He-Ne laser for 633 nm excitation. The average laser power was set at 14.1 mW. Renishaw WiRE 5.0 software was used for data retrieval and analysis. All Raman spectra, including the gelatin and water OH-stretch regions, were normalized to have the same intensity for the CH-stretch band at 2950 cm−1, representing the gelatin content in the samples. Additionally, the spectra containing the amide band were normalized to obtain the same intensity for the CH2 wag band at 1450 cm−1. All spectra were subjected to smoothing and baseline subtraction to eliminate the background signal.
8
Raman Analysis of Steel Corrosion Products
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Raman spectroscopy was employed as a powerful tool to discern the constituents of steel corrosion products. Through comparison of the obtained spectra with reference spectra from reputable literary sources, distinct phases of the corrosion product could be identified. The measurements were conducted using a Renishaw inVia Reflex confocal Raman microscope. For excitation, a 532 nm (Renishaw DPSSL laser, 50 mW) laser, filtered to 1% of its maximum intensity, was employed alongside an 1800 lines/mm diffraction grating. The Raman data obtained were then processed and analyzed using the Renishaw WiRE 5.3 software package. The spectral processing involved baseline subtraction, spectrum normalization, and curve fitting techniques to ensure accurate and reliable data interpretation.
9
Multimodal Analytical Toolkit Utilization
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10
Raman Spectroscopy of Material Samples
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Samples on stubs were analysed using a Renishaw inVia Qontor microscope with a 532 nm laser. Three points of interest were analysed to account for potential variability within the sample. Raman spectra were imported into the software Renishaw WiRE 5.3 and processed as follows: spikes were removed and the range 1100–1700 cm−1 was selected. Finally, the three spectra were smoothed and merged.
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