Artificially Generated Male-Specific Rabbit Antibody Against DBY-Multi-Epitope Fusion Protein And Its Immunoreactive Examination

DEAD (Asp-Glu-Ala-Asp) box polypeptide 3, Y-linked (DBY or Ddx3y) is a candidate gene for male-specific antigen. The DBY gene detected in capacitated mouse sperm codes putative ATP-dependent RNA helicase. The objective was to produce anti-predicted DBY multi-epitope fusion protein antibody, which co...

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Autores principales:	Wang Naidong, Tan Qinghui, Yuan Anwen, Xue Liqun
Formato:	article
Lenguaje:	EN
Publicado:	Sciendo 2015
Materias:	dby antibody spermatozoa gender selection Veterinary medicine SF600-1100
Acceso en línea:	https://doaj.org/article/fa7298c796b049bbb8cf8d95fdf4b2ef
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spelling	oai:doaj.org-article:fa7298c796b049bbb8cf8d95fdf4b2ef2021-11-17T21:27:50ZArtificially Generated Male-Specific Rabbit Antibody Against DBY-Multi-Epitope Fusion Protein And Its Immunoreactive Examination1820-744810.1515/acve-2015-0027https://doaj.org/article/fa7298c796b049bbb8cf8d95fdf4b2ef2015-09-01T00:00:00Zhttps://doi.org/10.1515/acve-2015-0027https://doaj.org/toc/1820-7448DEAD (Asp-Glu-Ala-Asp) box polypeptide 3, Y-linked (DBY or Ddx3y) is a candidate gene for male-specific antigen. The DBY gene detected in capacitated mouse sperm codes putative ATP-dependent RNA helicase. The objective was to produce anti-predicted DBY multi-epitope fusion protein antibody, which could be used to determine male specificity of DBY. Epitope prediction is to aid the design of molecules that can mimic structure and function of a genuine epitope, is a useful tool in protein molecule design. This study predicted the DBY epitopes, prepared rabbit poloclonal antibody against DBY multi-epitope fusion protein, then investigated its immunoreactivity. The fusion protein used as the antigen consisted of three regions of DBY with greatest divergence from other family members, cloned together in-frame (with a His tag to facilitate purification). The resulting antibody recognized both the DBY1-2-3 fusion protein and an endogenous DBY protein of the same size. Furthermore, DBY protein was present (Western blot) in testis, male mouse splenocytes and brain, whereas a weaker band was present in the female brain and splenocytes, and finally, ovary produced only a barely visible protein band. Optical density of DBY protein was higher for males versus corresponding tissues from females. Finally, positive signals of DBY1-2-3 antibody were present on only ~60% of mature murine sperm (based on immunofluorescent staining and flow cytometry), in accordance with the expected proportion of Y-bearing sperm. We hypothesized that our antibodies recognized a specific epitope present in subpopulations of mouse sperm. Therefore, we concluded that anti-DBY1-2-3 antibody could be an alternative way of producing antibodies to DBY protein. Furthermore, this novel DBY antibody against a multi-epitope artificial antigen has potential for both investigating male-specific binding of DBY and as a new method of sex selection.Wang NaidongTan QinghuiYuan AnwenXue LiqunSciendoarticledbyantibodyspermatozoagender selectionVeterinary medicineSF600-1100ENActa Veterinaria, Vol 65, Iss 3, Pp 328-338 (2015)
institution	DOAJ
collection	DOAJ
language	EN
topic	dby antibody spermatozoa gender selection Veterinary medicine SF600-1100
spellingShingle	dby antibody spermatozoa gender selection Veterinary medicine SF600-1100 Wang Naidong Tan Qinghui Yuan Anwen Xue Liqun Artificially Generated Male-Specific Rabbit Antibody Against DBY-Multi-Epitope Fusion Protein And Its Immunoreactive Examination
description	DEAD (Asp-Glu-Ala-Asp) box polypeptide 3, Y-linked (DBY or Ddx3y) is a candidate gene for male-specific antigen. The DBY gene detected in capacitated mouse sperm codes putative ATP-dependent RNA helicase. The objective was to produce anti-predicted DBY multi-epitope fusion protein antibody, which could be used to determine male specificity of DBY. Epitope prediction is to aid the design of molecules that can mimic structure and function of a genuine epitope, is a useful tool in protein molecule design. This study predicted the DBY epitopes, prepared rabbit poloclonal antibody against DBY multi-epitope fusion protein, then investigated its immunoreactivity. The fusion protein used as the antigen consisted of three regions of DBY with greatest divergence from other family members, cloned together in-frame (with a His tag to facilitate purification). The resulting antibody recognized both the DBY1-2-3 fusion protein and an endogenous DBY protein of the same size. Furthermore, DBY protein was present (Western blot) in testis, male mouse splenocytes and brain, whereas a weaker band was present in the female brain and splenocytes, and finally, ovary produced only a barely visible protein band. Optical density of DBY protein was higher for males versus corresponding tissues from females. Finally, positive signals of DBY1-2-3 antibody were present on only ~60% of mature murine sperm (based on immunofluorescent staining and flow cytometry), in accordance with the expected proportion of Y-bearing sperm. We hypothesized that our antibodies recognized a specific epitope present in subpopulations of mouse sperm. Therefore, we concluded that anti-DBY1-2-3 antibody could be an alternative way of producing antibodies to DBY protein. Furthermore, this novel DBY antibody against a multi-epitope artificial antigen has potential for both investigating male-specific binding of DBY and as a new method of sex selection.
format	article
author	Wang Naidong Tan Qinghui Yuan Anwen Xue Liqun
author_facet	Wang Naidong Tan Qinghui Yuan Anwen Xue Liqun
author_sort	Wang Naidong
title	Artificially Generated Male-Specific Rabbit Antibody Against DBY-Multi-Epitope Fusion Protein And Its Immunoreactive Examination
title_short	Artificially Generated Male-Specific Rabbit Antibody Against DBY-Multi-Epitope Fusion Protein And Its Immunoreactive Examination
title_full	Artificially Generated Male-Specific Rabbit Antibody Against DBY-Multi-Epitope Fusion Protein And Its Immunoreactive Examination
title_fullStr	Artificially Generated Male-Specific Rabbit Antibody Against DBY-Multi-Epitope Fusion Protein And Its Immunoreactive Examination
title_full_unstemmed	Artificially Generated Male-Specific Rabbit Antibody Against DBY-Multi-Epitope Fusion Protein And Its Immunoreactive Examination
title_sort	artificially generated male-specific rabbit antibody against dby-multi-epitope fusion protein and its immunoreactive examination
publisher	Sciendo
publishDate	2015
url	https://doaj.org/article/fa7298c796b049bbb8cf8d95fdf4b2ef
work_keys_str_mv	AT wangnaidong artificiallygeneratedmalespecificrabbitantibodyagainstdbymultiepitopefusionproteinanditsimmunoreactiveexamination AT tanqinghui artificiallygeneratedmalespecificrabbitantibodyagainstdbymultiepitopefusionproteinanditsimmunoreactiveexamination AT yuananwen artificiallygeneratedmalespecificrabbitantibodyagainstdbymultiepitopefusionproteinanditsimmunoreactiveexamination AT xueliqun artificiallygeneratedmalespecificrabbitantibodyagainstdbymultiepitopefusionproteinanditsimmunoreactiveexamination
_version_	1718425392353640448

Artificially Generated Male-Specific Rabbit Antibody Against DBY-Multi-Epitope Fusion Protein And Its Immunoreactive Examination

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