VGLUT1-, VGLUT2-, PSD95- and TH-immunofluorescence were analyzed in the NAc shell (Fig. 1b -from bregma 1.70 mm to 0.98 mm, according to Paxinos and Franklin, 2001 ). For analysis of VGLUT puncta levels, images (one per section, n = 6 per animal) were acquired on a confocal LSM 780 upright microscope (Zeiss) at 100× magnification (Plan-Apochromat 100×/1.46 Oil DIC M27; zoom 1.0; pixel size 0.83 μm) as 2 μm z-stacks (21 slices, 0.1 μm interval) and deconvolved using AutoQuant. Analysis of puncta was performed in ImageJ, using the functions ‘max intensity’ for stacks and ‘find maxima’ to determine puncta number and ‘colocalization threshold’ for colocalization analysis of VGLUT2 and TH puncta.
Lsm780 upright microscope
Manufactured by Zeiss
The LSM780 upright microscope is a high-performance confocal laser scanning microscope designed for advanced imaging applications. It offers a comprehensive set of features and capabilities to enable detailed analysis and visualization of biological samples. The LSM780 provides exceptional optical performance, advanced imaging techniques, and seamless integration with various imaging and analysis software.
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3 protocols using lsm780 upright microscope
1
Quantifying VGLUT and TH Puncta in NAc Shell
2
Confocal Imaging of Cellular Colocalization
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Confocal imaging was performed on a Zeiss LSM780 upright microscope of the Light Microscopy Facility, a core facility of the BIOTEC/CRTD at Technische Universität Dresden. Colocalization images were made with the C-Apochromat 63 × /1.20 W korr M27 objective. GFP and mRFP were simultaneously excited with 488 and 561-nm lasers, respectively and their fluorescences simultaneously detected in the range of 470 and 552 nm for GFP, and 575 and 650 nm for mRFP. Image analysis was performed with the Zeiss Zen2012 software (also used for acquisition) and the colocalization plugin JACoP (Bolte and Cordelières, 2006 (link)) of the Fiji software (Schindelin et al., 2012 (link)). For colocalization studies, 14-day-old seedlings (cultured on selective MS + 2% sucrose plates under long-day conditions) were imaged with water as imaging medium.
3
Quantifying Zebrafish Brain Mitosis
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