Journal: bioRxiv

Article Title: Shallow-angle intracranial cannula for repeated infusion and in vivo imaging with multiphoton microscopy

doi: 10.1101/2025.01.22.634409

Figure Lengend Snippet: Fluorophores were infused through the cannula system and multiphoton microscopy was used for in vivo imaging. Images are shown as maximum intensity projections of 3D volumes. Each fluorescent cell marker was delivered to n = 3 C57BL/6J mice. (a) Cell nuclei were labeled after infusion of Hoechst 33342. ( b ) Pericytes (green) were labeled after infusion of NeuroTrace 500/525 and Rhodamine B Dextran (red) was injected IV to act as an angiogram. ( c ) Astrocytes were labeled following the infusion of SR101. ( d ) Microglia (red) were labeled after infusion of Isolectin GS-IB4, Alexa Fluor 594 Conjugate. The vasculature was labeled using IV injection of Fluorescein Dextran (green). ( e ) Lysosomes were labeled after infusion of LysoTracker Green DND-26. ( f ) Perineuronal nets (green) were labeled after infusion of Fluorescein labeled WFL and Rhodamine B Dextran (red) was injected IV. ( g ) Fluorescent Aβ42 (green) was infused to demonstrate perivascular clearance out of the brain (n = 3 C57BL/6J mice). Rhodamine B Dextran was injected to label the vasculature (red). ( h ) ThioS was infused and showed labeling of amyloid plaques and CAA in APP/PS1 mice (n = 3). ( i ) Thiazine Red was infused in rTg4510 mice (n = 3) and showed labeling of neurofibrillary tangles (red). Fluorescein Dextran (green) was injected IV to serve as an angiogram.

Article Snippet: The following volumes and concentrations were used for labeling of various cell types and structures in the brain: 6 µL of 89.0 µM Hoechst 33342 for labeling cell nuclei, 1 µL of 1:5 dilution of stock NeuroTrace 500/525 for labeling pericytes, 1 µL of 20 µM SR101 for labeling astrocytes, 1 µL of 100 µg/ml Isolectin GS-IB4, Alexa Fluor 594 Conjugate for labeling microglia, 1 µL of 100 µM LysoTracker Green DND-26 for labeling lysosomes, 1 µL of 100 µg/ml Fluorescein labeled Wisteria Floribunda Lectin (Vector Laboratories) for labeling PNNs, 2 µL of 1 mg/ml HiLyte Fluor 488-labeled Aβ42 (Anaspec), 1 µL of 100 µg/ml ThioS (Sigma Aldrich) for labeling of amyloid plaques, 2 µL of 5 mg/ml Thiazine Red (Sigma Aldrich) for labeling of NFTs.

Techniques: Microscopy, In Vivo Imaging, Marker, Labeling, Injection, IV Injection