Aperfeiçoamento da obtenção e processamento de imagem por microtomografia síncrotron para estudo morfométrico cerebral da Drosophila melanogaster

Abstract

Synchrotron radiation phase-contrast microtomography (SR-PhC-µCT) has emerged as a powerful technique in biomedical science for non-destructive visualization of internal structures with very similar densities without the need for chemical staining and with high resolution. This approach eliminates the need for sample preparations that could potentially damage micro and nanoscale structures. In this work, we utilized the SYRMEP beamline at Elettra synchrotron in Italy to investigate the internal structures of Drosophila melanogaster, commonly known as the fruit fly, renowned as an ideal model for studying neurodegenerative diseases due to its genetic correspondence with human disease-causing genes. We employed a single distance phase-contrast regimen combined with phase retrieval techniques pioneered by Paganin (2002). The samples were measured in two configurations, one with an average energy of 16.7 keV and the other with 25 keV, both using a filtered polychromatic X-ray beam and a 16-bit air/water-cooled sCMOS camera in combination withappropriate optics, obtaining pixel sizes of 1.5 and 0.9 μm. The samples, mounted on Pluronic hydrogel (F-127) and 100% ethanol for stabilization, were positioned at two sample-detector distances, one of 100 mm and the other of 150 mm. 1200 and 1800 projections around 180° were acquired. Different parameters for phase retrieval have been tested and evaluated to achieve optimal contrast between the brain and neighboring structures. Reconstruction was performed using the filtered back projection (FBP) algorithm, with careful parameter selection to guarantee the best contrast while minimizing information loss. Our results demonstrate the effectiveness of SR-PhC-µCT in visualizing and differentiating the fly's brain, as well as allowing for a precise quantification of the brain’s volume. Looking forward, the potential of utilizing advanced synchrotron sources like the MOGNO beamline at Sirius (Brazil) presents exciting prospects.