The corneal tissue sections were permeabilized with 0.5% Triton X-100, then blocked with goat serum (Solarbio, China) for 60 min. anti-NLRP3 and anti-CD80 (Santa Cruz Biotechnology, USA) were prepared at a dilution ratio of 1:100 and incubated with the cornea sections at 4 °C overnight. After incubating with the secondary antibodies at room temperature for 1 h, an anti-fade solution (containig DAPI) (Solarbio) was added, and the sections were observed under a uorescence microscope.
Cell immuno uorescence staining was carried out by counting and inoculating the THP-1 cells in a confocal culture dish. Paraformaldehyde (4%) was added, and the cells were xed for 15 minutes. The remaining steps were the same as described above for immuno uorescence staining of the cornea.
To observe the induction of autophagy, autophagosome-lysosome living cell dye DALGreen (Dojindo) was added to the macrophages cultured in the confocal dish for 30 min according to the manuscript. The remaining steps were the same as above for the cornea and cell immuno uorescence methods. Relative uorescence intensity was analyzed by ImageJ software.
Cell immuno uorescence staining was carried out by counting and inoculating the THP-1 cells in a confocal culture dish. Paraformaldehyde (4%) was added, and the cells were xed for 15 minutes. The remaining steps were the same as described above for immuno uorescence staining of the cornea.
To observe the induction of autophagy, autophagosome-lysosome living cell dye DALGreen (Dojindo) was added to the macrophages cultured in the confocal dish for 30 min according to the manuscript. The remaining steps were the same as above for the cornea and cell immuno uorescence methods. Relative uorescence intensity was analyzed by ImageJ software.