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. 2012 Jun 19;7(6):e39397. doi: 10.1371/journal.pone.0039397

Figure 3. Evaluation of the intracellular ROS in zebrafish spermatozoa during cryopreservation and its effect on viability and motility.

Figure 3

(A) Overall, cryopreservation decreased the viability of zebrafish sperm by approximately 50%, but the addition of CAT to the methanol improved its post-thaw viability 16% (P<0.05, ANOVA and Tukey’s Multiple Comparison test, F = 60.5). (B) Cryopreservation increased ROS over 2.7-fold (P<0.05, ANOVA, and Tukey’s Multiple Comparison test F = 6.2), but the addition of CAT did not mitigate it (P>0.5, ANOVA). (C) Motility demonstrated the greatest sensitivity in our treatments. Prior to cryopreservation, the 8% methanol treatment reduced motility by ∼50%, and the addition of CAT to the 8% methanol returned the motility to control levels. In parallel, the cryopreserved sperm (8% methanol) decreased the pot-thaw motility of zebrafish sperm from 83% (control) to 17%, however the addition of CAT doubled this post-thaw motility (P<0.05, ANOVA and Tukey’s Multiple Comparison test, F = 22.6). Bars with the same letter are not significantly different (P>0.05), but bars with different letters are (P<0.05). X- axis categories and sample sizes were: Control =  live sperm stained with DHE, N = 13; CAT (200 U/ml, concentration used in all CAT treatments), N = 9; 8% Methanol, N = 12; 8% Methanol + CAT 200, N = 12; Cryo 8% Methanol  =  sperm cryopreserved according to [6] with 8% methanol, N = 13; Cryo 8% Methanol + CAT, N = 13.