Objective To investigate the outcomes of fixation of rat retina paraffin sections with three different methods. Methods Nine SD rats were randomly divided into group A (retina fixation with improved solution, n=3), group B (retina fixation with 4% paraformaldehyde, n=3) and group C (retina fixation with cardiac perfusion of 4% paraformaldehyde, n=3). Paraffin sections were prepared 24 h after retina fixation, HE staining was performed, and the outcomes of fixation were compared. Results There was no eyecup deformation in group A,  a few eyecup deformations in group B and few eyecup deformation in group C. It was observed by microscopy that each layer of the retina structure was intact in group A,  and there was retinal detachment in group B and group C. Conclusion Eyeball fixation with improved fixation solution may result in better outcomes than fixation with 4% paraformaldehyde or fixation with cardiac perfusion, and is suitable for further research with retinal paraffin sections.