Currently, testicular sperm extraction is successfully combined with intracytoplasmic sperm injection into the oocyte (ICSI). Several pieces of a testicular biopsy can be frozen and thawed until the ICSI attempt. In this study, the effects of freezing-thawing on the morphology of rat testicular biopsies stored in different cryopreservation media were analysed. Each cryopreservation medium contained glycerol and/or dimethyl sulfoxide (DMSO) as cryoprotectants. In general, both glycerol and DMSO, when applied at moderate concentrations (6-25%), preserved the structure of the seminiferous epithelium. The freezing-thawing procedure had no significant effect on tubular diameter; however, it caused a 'folding' of the lamina propria and notable damage to Sertoli cells, spermatogonia and spermatocytes. Round and elongated spermatids and spermatozoa displayed occasional nuclear damage, vacuolization, and shrinkage/swelling of the cytoplasm. However, the vast majority of these cells maintained their normal structure in nearly all the applied cryomedia. It is concluded that freezing-thawing of testicular biopsies, and the cryopreservation medium, have a significant impact on the structure of the seminiferous epithelium, particularly on its basal compartment.