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This study has established a successful protocol to cryopreserve lumpfish Cyclopterus lumpus (Linnaeus, 1758) milt. Three cryosolutions were tested based on Mounib’s medium; the original medium including reduced l-glutathione (L-Glu), the basic sucrose and potassium bicarbonate medium without L-Glu, or with hen’s egg yolk (EY). Dimethyl sulphoxide (DMSO) was used as the cryoprotectant along with all three diluents in a 1 to 2 dilution. Cryopreservation was performed with the mentioned cryosolutions at two freezing rates. Motility percentages of spermatozoa were evaluated using ImageJ with a computer assisted sperm analyzer (CASA) plug-in. Findings revealed that spermatozoa cryopreserved in Mounib’s medium without L-Glu had a post-thaw motility score of 8.64 percentage points (pp) higher than to that with added L-Glu, and an addition of EY to the Mounib’s medium lowered the post-thaw motility score by 15.07 pp. The difference in motility between both freezing rates was 5.23 pp, and samples cryopreserved on a 4.8 cm high tray resulted in a better post-thaw motility score. Cryopreserved milt had a 23.76 pp lower post-thaw motility score when compared with fresh milt. Cryopreservation of lumpfish milt has, to our knowledge, never been successfully carried out before. The established protocol will be a main contributing factor in a stable production of lumpfish juveniles in future.