| Autor: |
Kyle T. Nguyen, Alexandre R. Sathler, Alvaro G. Estevez, Isabelle E. Logan, Maria Clara Franco |
| Jazyk: |
angličtina |
| Rok vydání: |
2024 |
| Předmět: |
|
| Zdroj: |
Frontiers in Cell and Developmental Biology, Vol 12 (2024) |
| Druh dokumentu: |
article |
| ISSN: |
2296-634X |
| DOI: |
10.3389/fcell.2024.1420161 |
| Popis: |
A common problem in confocal microscopy is the decrease in intensity of excitation light and emission signal from fluorophores as they travel through 3D specimens, resulting in decreased signal detected as a function of depth. Here, we report a visualization program compatible with widely used fluorophores in cell biology to facilitate image interpretation of differential protein disposition in 3D specimens. Glioblastoma cell clusters were fluorescently labeled for mitochondrial complex I (COXI), P2X7 receptor (P2X7R), β-Actin, Ki-67, and DAPI. Each cell cluster was imaged using a laser scanning confocal microscope. We observed up to ∼70% loss in fluorescence signal across the depth in Z-stacks. This progressive underrepresentation of fluorescence intensity as the focal plane deepens hinders an accurate representation of signal location within a 3D structure. To address these challenges, we developed ProDiVis: a program that adjusts apparent fluorescent signals by normalizing one fluorescent signal to a reference signal at each focal plane. ProDiVis serves as a free and accessible, unbiased visualization tool to use in conjunction with fluorescence microscopy images and imaging software. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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