Serpa Carangui, Erika MargothSamaniego Campoverde, Jorge XavierGalarza Lucero, Diego AndrésSoria Parra, Manuel ElíasOchoa Guillén, Juan DiegoÁlvarez Palacios, David AlejandroDe la Cruz Tandazo, Francisco Javier2022-12-022022-12-0220220000-00000936-6768, e1439-0531http://dspace.ucuenca.edu.ec/handle/123456789/40411https://onlinelibrary.wiley.com/toc/14390531/2022/57/S5This study evaluated the effect of penetrating cryoprotectant agents (CPA) and the cryosurvival of three freezing protocols on the kinematics and integrity of membranes of frozen-thawed stallion sperm. Twenty-four ejaculates of four adult Arabian horses were collected in six weekly sessions (six ejaculates/horse). Each ejaculate was divided into two aliquots. With the first aliquot, three CPA treatments were conformed: 5% glycerol (GLY), 5% dimethylformamide (DMF), and 3%–3% DMF–GLY combination, and the sperm samples were frozen exposing them to liquid nitrogen (LN2) vapors. The second aliquot was diluted with freezing medium plus 5% DMF and the sperm samples were frozen in three freezing protocols: (P1) Styrofoam cryo-box (30 × 29 × 31 cm of length, width, and height, respectively) with two ramps (at 17 and 7 cm above LN2); (P2) freezing unit® (Minitüb, Germany); and (P3) programmable TK 4000-freezer® (Compacta, Brazil). The DMF-GLY combination and DMF yielded higher (p<.05) post-thaw values than the GLY regarding the motility (SM: 54.2±2.25 and 50.2±1.80 vs. 41.4±2.35%, respectively), curvilinear velocity (VCL: 58.0±1.71 and 54.0±1.58 vs. 42.3±1.60 µm/s), and the proportion of sperm with intact plasma and intact acrosome (IPIA: 58.0±1.11 and 52.6±0.99 vs. 42.5±1.07%). Furthermore, the P1 protocol produced a similar (p>.05) post-thaw SM, VCL, and IPIA than the other protocols. Indeed, the P1 and P3 protocols yielded lower proportion (p<.05) of sperm with damaged plasma and damaged acrosome than the P2 protocol after thawing (3.7±0.18 and 3.1±0.18 vs. 6.1±0.44%, respectively). In conclusion, the addition of DMF or combined with GLY to freezing medium, and the freezing with Styrofoam cryo-box with two ramps increase the cryosurvival of Arabian stallion spermatozoa.es-ESFreezing protocolsDimethylformamideSpermatozoaArabian stallionGlycerolARTÍCULO DE CONFERENCIA10.1111/rda.14227