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[Preparation of SUN5-specific polyclonal antibody for detection of SUN5 expression in human germ cells]
Jing-Fang Wang1, Li-Hua Huang, Yong Wu
1Center for Experimental Medicine Research, Third Xiangya Hospital, Central South University, Changsha 410013, China.E-mail: foreveryouth@yeah.net.
Objective:
To prepare a specific polyclonal antibody against full-length SUN5 for detecting the expression of SUN5 in human germ cells.
Methods:
Bioinformatic methods were used to compare the full-length SUN5 and its variant SUN5β, and a short peptide was designed based on the differential region to prepare SUN5 antibody. The prepared antibody was used to detect the expression of SUN5 in Ntera-2 cells and in human germ cells by Western blotting and immunofluorescence assay.
Results:
The short peptide was correctly synthesized and SUN5 antibody was obtained and purified. Western blotting showed that the prepared antibody was capable of recognizing full-length SUN5 in Ntera-2 cells, and SUN5 expression was localized on the nuclear membrane and in the cytoplasm as shown by immunofluorescence assay. Using this antibody, we detected SUN5 expression in the spermatocytes, round spermatids and sperms in human germ cells.
Conclusion:
We successfully prepared SUN5-specific antibody. SUN5 is expressed in the spermatocytes, round spermatids and sperms in human germ cells, suggesting its important role in spermatogenesis.
