Reflex raman microscope

Resonance Raman spectra of cytochromes were recorded with a Renishaw inVia Reflex Raman microscope with a ×50 objective. A 532 nm laser was used with 5 s exposure at 50% laser power for all measurements. At least 10 individual spectra were recorded from the middle of the filaments to obtain an averaged spectrum. Spectra were subsequently corrected for background scattering from the glass slide and medium by recording additional spectra next to the filaments. Reduced spectra were measured in 10 mM sodium dithionite in artificial seawater (ASW) and oxidized spectra were measured in air-saturated ASW.

Raman scattering measurements were performed with an inVia Renishaw Reflex Raman Microscope in micro-Raman mode, with an edge filter with spectral cutoff at 80 cm⁻¹ and a 2400 grooves/mm grating for the analysis of the scattered light. A frequency doubled Nd:YAG laser (532 nm) was used as excitation source. The laser beam was focused through a microscope objective with magnification ×100L and numerical aperture 0.85, which results in a spot size of about 0.76 µm. Laser power was not higher than 0.44 mW. For each fluence, 2–4 measurements were done in different locations of every crater created by ion irradiation (Figure 8). For deconvolution of the Raman spectra, the method by Ribeiro-Soares et al. has been used [58 (link)]. The peaks for the D band at 1345 cm⁻¹, the G band at 1585 cm⁻¹ and the D’ band at 1615 cm⁻¹ have been fitted by Lorentzian functions while Gaussian functions have been used to fit the contribution from highly disordered areas close to 1250 cm⁻¹ and 1490 cm⁻¹. In order for the fitting algorithm to converge, the peak positions of the Gaussian peak at 1250 cm⁻¹ and the D’ peak needed to be fixed. After fitting, integrated peak intensities of the D and G bands and the full width at half maximum (FWHM) of the G and D’ bands have been extracted from every Raman spectra. Error bars have been created using the standard deviation of the different parameters.