Separation and purification of human testicle spermatogonia with Percoll centrifugation combined with different time of cellular adhesiveness

Objective To research the effective methods of separation and purification of human testicle spermatogonia.Methods The two-step enzymatic digestion was used to obtain germ cells suspension to be centrifuged by Percoll discontinuous density gradient centrifugation.Finally,purified human spermatogonias were obtained according to different time and times of cellular adhesiveness(per 2,3,4 h and one or twp times).Results ①The percentage of spermatogonia only after separated by Percoll was 48.5%,and that of purified one time by interval 2,3,4 h was 51.3%,59.0%,56.4% respectively;②The percentage of spermatogonia was 57.7%,67.8%,57.8% respectively through 2,3,4 h interval and continuous 2 times;The highest was in interval of 3 h;③Compared with purified one time,that in interval of 2,3 h and two times were higher than that of one time;But that of 4 h and two times had no difference from that of one time.Conclusions ①Percoll centrifugation combined with different time of cellular adhesiveness can effectively separate and purify the human spermatogonia.②The percentage of spermatogonia of purification by two times is higher than that of one time by interval of 2,3 h;③If purification by continuous two times,interval 3 h is most effective.