Lsm 700 axioimager

All litters from C57BL/6J mice were randomly divided into two groups: vehicle- and polyI:C-treated. Neonatal C57BL/6J mice were administered with a daily subcutaneous injection of saline (control) or polyI:C (5 mg/kg, Sigma-Aldrich) between PD2 and PD6. Immunohistochemistry was conducted as described previously [17 (link)]. Neonatal mice were deeply anesthetized with diethyl ether 24 h after the final polyI:C treatment and perfused transcardially with saline, followed by 4% paraformaldehyde in phosphate-buffered saline (PBS, pH 7.4). The brains were removed and cryoprotected. 20-μm-thick coronal brain sections were cut on a cryostat and mounted on slides. The sections were denatured in a microwave oven in 0.01 M citrate buffer (pH 6.0). After blocking with 5% donkey and 5% goat serum/PBS, mouse anti-glial fibrillary acidic protein (GFAP, a marker for astrocytes, 1:1,000, Sigma-Aldrich) and rat anti-Fstl1 (1:100, R&D systems) were added to the sections. After washing in PBS, goat anti-mouse Alexa Fluor (AF) 568 and anti-rat AF488 antibodies (1:1,000, Invitrogen) were added to the sections. The samples were observed using a confocal-laser scanning microscope (LSM 700 Axio Imager; Zeiss).

3rd instar larval eye discs were stained in 5 µM SYTO Red Fluorescent Nucleic Acid Stain (Invitrogen) and imaged on a confocal microscope LSM 700/Axioimager (Zeiss).