This study aimed to examine the effects of the interaction of the addition of vitamin C, the type of diluent, and the duration of storage at 5°C on the quality and fertility of Rooster Kampung spermatozoa and the fertile period of Red Kedu hen eggs. Semen was collected from 10 roosters subjected to vitamin C level treatment (0 mg, 0.1 mg, 0.2 mg, or 0.3 mg), two diluent types (lactated Ringer (RL) and skim milk + 50 mM glucose), and storage durations (0 hours, 3 hours, 6 hours, or 9 hours). The sperm samples meeting the quality standards were inseminated into 64 Red Kedu hens. Phase 1 of the study used a factorial randomised design (2´4´4) followed by variance analysis and orthogonal polynomial testing. Phase 2 applied the eight best result treatments from phase 1 via a completely randomised design with Tukey’s test for post hoc analysis. The results revealed a highly significant interaction effect (P<0,01) between vitamin C level and diluent type on motility, plasma membrane integrity (IPM), and sperm abnormality. The interaction effect of vitamin C level and storage duration had a very significant effect (P<0.01) on motility, IPM, and viability. The interaction effect of diluent type and storage duration significantly affected (P<0.01) motility and IPM. However, no significant effect (P>0.05) among all three variables was observed on fertility or the fertile period. In summary, the addition of 0.3 mg of vitamin C, lactated Ringer’s diluent, and 6 hours of storage at 5°C maintained sperm quality and resulted in 51.78±37.64 fertility and a fertile period of 15 days in Red Kedu hens.