Addition of cholesterol-loaded cyclodextrin (CLC) to the diluents of mammalian semen increased stability and rigidity of phospholipid hydrocarbon chains of plasma membrane during sperm cryopreservation process. CLC has been tested successfully as cryoprotectant in various livestock sperm cryopreservation protocols but its efficacy for cryopreserving of fish sperm has not previously been tested. In the present study, different cholesterol loaded cyclodextrin concentrations were evaluated for the cryopreservation of carp (Cyprinus carpio) sperm. Sexually mature fish were induced to spermiation and ovulation with Ovopel. The extenders were prepared by using 300mM glucose and 10% DMSO supplemented with different concentrations of CLC (0.5, 1.0, 1.5, 2.0, 2.5, and 3.0mg per 120×106 spermatozoa) and without CLC (control). The pooled semen was diluted separately at a ratio of 1:3 (v/v) by using CLC extenders. Diluted semen placed into 0.25ml straws were equilibrated at 4°C for 15min and frozen in liquid nitrogen vapor. Fertilization was conducted using a ratio of 1×105 spermatozoa/egg.Fresh sperm with no treatment showed the greatest sperm motility, duration of motility, viability, and fertilization results compared to the other tested cryopreserved and control groups (p<0.05). Supplementation of 1.5mg CLC to the extender showed the best cryoprotective effect for sperm motility, duration of motility, and viability against freezing damage in comparison to extenders containing 2.5mg, 3.0mg CLC, and control group (p<0.05). Cryopreserved sperm containing 1.5mg CLC provided greater result in term of fertilization success when compared to other extenders containing 0.5, 2.5, and 3.0mg CLC or control (p<0.05). The amount of CLC effected post-thaw sperm quality and fertility as a dose-dependent manner.It is concluded that treatment of cholesterol-loaded cyclodextrin for carp sperm cryopreservation significantly improves cell cryosurvival and fertilization.