Grass Carp Reovirus Infection Assay

GCO cells were cultured in M199 medium (HyClone, USA) supplemented with 10% fetal bovine serum (FBS), 100 IU/mL of penicillin, and 100 mg/mL of streptomycin under humidified conditions with 5% CO₂ at 28°C. Grass carp reovirus, which was isolated and identified as subtype I by our laboratory (50 (link)), was used in the study of viral infection. Rabbit polyclonal antibodies against the GCRV nonstructural protein NS80 (anti-NS80) and structural protein VP5 (anti-VP5) were prepared in our laboratory (51 (link)). Mouse anti-double-stranded RNA (J2 clone) antibody was purchased from Nordic-MUbio (Susteren, Netherlands). Horseradish peroxidase (HRP)-conjugated goat anti-rabbit IgG was purchased from Sigma-Aldrich (St. Louis, MO, USA). Alexa Fluor 594-conjugated goat anti-rabbit IgG (H+L) was purchased from Cell Signaling Technology (Danvers, MA, USA). Alexa Fluor 594-conjugated goat anti-mouse IgG (H+L) was purchased from Abcam (Cambridge, UK). The LD probes oil red O and Bodipy 493/503 were purchased from Beyotime (Shanghai, China) and GLPBIO (CA, USA). C75, TOFA, IBMX, palmitic acid (PA), atglistatin, and CAY10499 were purchased from GLPBIO (CA, USA). Triacsin C and isoproterenol were purchased from Abcam and MedChemExpress (USA).