Retinal Immune Cell Profiling in Mice

The eyeballs were enucleated from C57 and rd1 mice at P14, P21, P28, and P180. Retinae were dissected out and placed in 1 ml RPMI with a pH of 7.4 regulated by Hepes (1:100, MP BIOMEDICALS), and disaggregated by gently pipetting up and down through a wide bore pipette tip as previously described (Noailles et al., 2016 (link)). This cell suspension was filtered through a 70-μm strainer (BD Biosciences, San Diego, CA, United States) to prevent cell clumps. The cells were washed with phosphate-buffered saline (PBS) containing 1% FBS (Life Technologies, Grand Island, NY, United States) and 1% Hepes (MP BIOMEDICALS), and were stained by CD45 (2 ug/ml, BD pharmingen^TM), CD11b (5 ug/ml, eBioscience), CD86 (1 ug/ml, BD pharmingen^TM), and CD206 (5 ug/ml, Biolegend) antibodies for 30 min at room temperature. A four-laser Becton-Dickinson FACS Calibur (BD Biosciences) was used to collect the data, and FlowJo software was used for analysis.