Background and Aim: To enhance the reproductive potential and increase productivity and population of cows, spermatozoa sex-sorting technology is required. This study aimed to examine the effect of sexing sperm, separated using a bovine serum albumin (BSA) column with varying incubation durations and centrifugation methods, for successful artificial insemination.

Materials and Methods: Six Simmental bulls and 30 cows (n = 30) as the recipients were selected for this study at Balai Pembibitaan Hijauan Pakan Ternak Sembawa Indonesia. The study parameters included sperm motility, viability, plasma membrane integrity, and conception rate (CR). The experiment was divided into three protocols to find out differences in some parameters: (1) BSA incubation time effect (P) with P1 (40 min), P2 (50 min), and P3 (60 min); (2) freezing time effect with before freezing and after-thawing treatments; and (3) CR determined by measuring the proportion of pregnant cows following insemination with non-sexed, X-bearing, and Y-bearing sperms without centrifugation (n = 15) (A0, A1, and A2) and with centrifugation (n = 15) (B0, B1, and B2) in the acquired data, which were counted using the Statistical Package for the Social Sciences version 21 program. Analysis of variance was utilized to evaluate all treatments at various levels.

Results: The results demonstrated that centrifugation time influenced all sperm quality metrics for sperm containing X and Y (p < 0.05). The non-return rate (NRR) of non-sexed frozen semen, both centrifuged (A0) and not centrifuged (B0), was more significant than frozen semen produced by sexing X and Y spermatozoa. The NRR indicated a value of 80% based on the number of lactating cows.

Conclusion: Bovine serum albumin incubation and centrifugation protocols influenced and decreased all sperm quality indicators throughout the sexing procedure and could still be used as a sexing protocol. Furthermore, regarding NRR and service per conception, non-sexual treatment is superior to sexing treatment.

Keywords: bovine serum albumin, centrifugated, conception rate, incubation, sexing, sperm.