AstroDot

AstroDot analyzes the spatial distribution of messenger RNA (mRNA) within GFAP-immunolabeled astrocytes to identify and quantify mRNA localization in somata, large processes, and fine processes.

Key Features:

• Three-Dimensional Histological Methodology: Integrates three-dimensional confocal imaging with in situ hybridization and GFAP immunostaining to map mRNA within astrocytic compartments.
• Custom ImageJ Plug-in: Implements a custom ImageJ plug-in for identification and quantification of mRNAs within GFAP-labeled somata, large processes, and fine processes.
• Integration with AstroStat R Package: Exports quantified mRNA distribution data for downstream statistical and analytical processing using the AstroStat R package.

Scientific Applications:

• Characterization of GFAP Isoforms: Localizes mRNAs encoding GFAP-α and GFAP-δ and demonstrates isoform-enriched distributions across hippocampal regions and relative to amyloid plaques in the APPswe/PS1dE9 mouse model.
• Validation Across Cell Types: Detects ubiquitous Rpl4 mRNA in astrocyte and microglia processes and confirms mRNA detection in Iba1-immunolabeled microglia.

Methodology:

Three-dimensional confocal imaging combined with in situ hybridization and GFAP immunostaining, image-based identification and quantification via a custom ImageJ plug-in, and downstream data analysis using the AstroStat R package.