Abstract: Objective To investigate the expression of CD86 co-stimulatory molecules in rejected rat corneal graft in situ after penetrating keratoplasty. Methods Rat models of orthotopical corneal transplantation were established, and after the occurrence of graft rejection, immunohistochemical staining was performed on the corneal whole-mounts and the spleen tissue of the recipient rats. Results CD86⁺ cells were detected in the epithelium of the corneal graft during allograft rejection but not in normal cornea. Conclusion In situ expression of costimulatory molecules in the corneal allograft may be related to the acute immune rejection after penetrating keratoplasty.