Anticorpo monoclonal contra uma proteína macho-específica de 19 kDa em espermatozóides bovinos: uma metodologia promissora para imunosexagem

Souza, Clóvis José Pascarelli; Matta, Marcos Fernandes Resende da; Cruz, Gilson Mendes; Alves, Elias Walter; Kanashiro, Milton Massaricho; Silva, José Frederico Straggiotti

doi:10.1590/S1516-35981999000100010

Resumos

Neste trabalho, foi purificada uma proteína macho específica com peso molecular de 19 Kda, a partir de espermatozóides de bovinos, por intermédio de cromatografia de gel filtração. O anticorpo monoclonal (Mab) desenvolvido (C11F) contra esta proteína discriminou duas populações de espermatozóides, como revelado por imunofluorescência em microscopia e citometria de fluxo. O epitopo reconhecido pelo Mab C11F esteve presente em 49,7% da população de espermatozóides. Quando adicionado ao sêmen bovino fresco, o Mab C11F reduziu a motilidade e promoveu a aglutinação dos espermatozóides.

espermatozóides; bovinos; imunosexagem; proteína macho específica; ac. monoclonal

In this work, a 19KDa, male specific protein, from bovine spermatozoa, was purified using gel filtration chromatography. A monoclonal antibody (Mab) developed (C11F) against this protein discriminated two populations of spermatozoa, as revealed by immunofluorescence in microscopy and flow citometry. The epitope recognized by Mab C11F was present in 49.7% of spermatozoa population. When added to fresh bovine semen, the Mab C11F reduced the motility and promoted agglutination of the spermatozoa.

spermatozoa; bovines; immunosexing; male specific protein; monoclonal antibody

MELHORAMENTO, GENÉTICA E REPRODUÇÃO

Anticorpo monoclonal contra uma proteína macho-específica de 19 kDa em espermatozóides bovinos: uma metodologia promissora para imunosexagem

Monoclonal antibody against male-specific protein of 19 KDa from bovine spermatozoa: a successful methodology for imunosexing

Clóvis José Pascarelli Souza^I,V; Marcos Fernandes Resende da Matta^I; Gilson Mendes Cruz^I,IV; Elias Walter Alves^II; Milton Massaricho Kanashiro^III; José Frederico Straggiotti Silva^I

^ILMGA/Universidade Estadual do Norte Fluminense - Campos

^IILQFPP e DBM - ICB/Universidade Federal do Rio de Janeiro

^IIILBR/Universidade Estadual do Norte Fluminense

^IVIESES/VFac. Veterinária e Zootecnia de Castelo - Castelo/ES

^VFIOCRUZ - RJ

RESUMO

Neste trabalho, foi purificada uma proteína macho específica com peso molecular de 19 Kda, a partir de espermatozóides de bovinos, por intermédio de cromatografia de gel filtração. O anticorpo monoclonal (Mab) desenvolvido (C11F) contra esta proteína discriminou duas populações de espermatozóides, como revelado por imunofluorescência em microscopia e citometria de fluxo. O epitopo reconhecido pelo Mab C11F esteve presente em 49,7% da população de espermatozóides. Quando adicionado ao sêmen bovino fresco, o Mab C11F reduziu a motilidade e promoveu a aglutinação dos espermatozóides.

Palavras-chave: espermatozóides, bovinos, imunosexagem, proteína macho específica, ac. monoclonal.

ABSTRACT

In this work, a 19KDa, male specific protein, from bovine spermatozoa, was purified using gel filtration chromatography. A monoclonal antibody (Mab) developed (C11F) against this protein discriminated two populations of spermatozoa, as revealed by immunofluorescence in microscopy and flow citometry. The epitope recognized by Mab C11F was present in 49.7% of spermatozoa population. When added to fresh bovine semen, the Mab C11F reduced the motility and promoted agglutination of the spermatozoa.

Key Words: spermatozoa, bovines, immunosexing, male specific protein, monoclonal antibody.

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Recebido em: 13/06/97

Aceito em: 28/08/98

BAHATTACHARYA, B.C., BANGHAM, A.D., KCRO, R.J. et al. 1966. An attempt to predetermine the sex of calves by artificial insemination whith spermatozoa separeted by sedimentation. Nature, 211:863.
BONDIOLI, K.R. 1992. Embryo sexing: a review of current techniques and their potential for commercial application in livestock production. J. Anim. Sci., 70:19-29.
BOZZO, S., RETAMAL, C.A. 1991. Geles unidimensionales - Um nuevo método densitométrico para computadores personales. Arch. Biol. Med. Exp, 24:181.
BRADLEY, M.P. 1989. Immunological Sexing of Mammalian Semen: Current Status and Future Options. J. Dairy Sci 72:3372-3380.
COLIGAN, J.E., Kruisbeek, A.M., MARGULIES, D.H. et al. 1991. Antibody Detection and Preparation. In: Currents Protocols in ImmunologyI, Jonh Wiley e Sons (editor), New York, V.1 capit. 2 part. 2 pg 2.5.1-2.6.6.
GOLDBERG, H.G., BOYSE, E.A., BENNETT, D. 1971. et al. Serological demonstration of H-Y (male) antigen on mouse sperm. Nature, 232:478-480.
HAFS, H. D., BOYD, L.J. 1971. Galvanic separation of X- and Y- chromosome bearing sperm. In Sex Ratio at birth-prospects for control (C. A. Kiddy and H. D. Hafs, eds.). Am. Soc. for Anim. Sci., p.85-97.
HAM, T.L., FLAHERTY, S.P., FORD, J.H. et al. 1993. Detection of X- and Y- bearing human spermatozoa after motile sperm isolation by swim-up. Fertility and Sterility, 60(6):1046-1051.
JOHNSON, L.A. 1991. Sex preselection in swine: altered sex ratios in offspring following surgical insemination of flow sorted X- and Y-bearing sper. Reprod. Dom. Animal, 26:309-314.
JOHNSON, L. A. 1992. Gender preselection in domestic animals using flow cytometrically sorted sperm. J. Anim. Sci, v.70, suppl.2, p. 8-18.
KOO, G.C., STACHPOLE, C.W., BOYSE, E.A. et al. 1973. Topographical location of H-Y antigen on mouse spermatozoa by immunoelectronmicroscopy. Proc. Nat. Acad. Sci., 70:1502-1505.
MEISTRICH, M. L. 1982. Potential and limitation of physical methods for separation of sperm bearing an X- or Y- chromosome. In: Prospects for sexing mammalian sperm (R.P. Amann and G. E. Seidel eds.), Colorado Associated University Press, Boulder, Colorado, p. 157-168.
MORUZZI, J.F. 1979. Selecting a mammalian species for the separation of X- and Y-chromosome-bearing spermatozoa. J. Reprod. Fertil., 37:319-323.
PETER, A. J., JONES, P.P., ROBINSON, J.P. 1993. Fraction of bovine spermatozoa for sex selection: A rapid immunomagnetic technique to remove spermatozoa that contain the H-Y antigen. Theriogenology, 40:1177-1185.
UTSUMI, K., HAYASHI, M., TAKAKURA, R. et al. 1993. Embryo sex selection by a rat male-specific antibody and the cytogenetic and developmental confirmation in cattle embryos. Mol. Reprod. and Devel., 34:25-32.
VAN VLECK, L.D., EVERETT, R. W. 1975. Genetic value of sexed semen to produce dairy heifers. J. Dairy Sci., 59(10):1802-1808.

Datas de Publicação

Publicação nesta coleção
02 Out 2013
Data do Fascículo
Fev 1999

Histórico

Aceito
28 Ago 1998
Recebido
13 Jun 1997

This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

[1] BAHATTACHARYA, B.C., BANGHAM, A.D., KCRO, R.J. et al. 1966. An attempt to predetermine the sex of calves by artificial insemination whith spermatozoa separeted by sedimentation. Nature, 211:863.

[2] BONDIOLI, K.R. 1992. Embryo sexing: a review of current techniques and their potential for commercial application in livestock production. J. Anim. Sci., 70:19-29.

[3] BOZZO, S., RETAMAL, C.A. 1991. Geles unidimensionales - Um nuevo método densitométrico para computadores personales. Arch. Biol. Med. Exp, 24:181.

[4] BRADLEY, M.P. 1989. Immunological Sexing of Mammalian Semen: Current Status and Future Options. J. Dairy Sci 72:3372-3380.

[5] COLIGAN, J.E., Kruisbeek, A.M., MARGULIES, D.H. et al. 1991. Antibody Detection and Preparation. In: Currents Protocols in ImmunologyI, Jonh Wiley e Sons (editor), New York, V.1 capit. 2 part. 2 pg 2.5.1-2.6.6.

[6] GOLDBERG, H.G., BOYSE, E.A., BENNETT, D. 1971. et al. Serological demonstration of H-Y (male) antigen on mouse sperm. Nature, 232:478-480.

[7] HAFS, H. D., BOYD, L.J. 1971. Galvanic separation of X- and Y- chromosome bearing sperm. In Sex Ratio at birth-prospects for control (C. A. Kiddy and H. D. Hafs, eds.). Am. Soc. for Anim. Sci., p.85-97.

[8] HAM, T.L., FLAHERTY, S.P., FORD, J.H. et al. 1993. Detection of X- and Y- bearing human spermatozoa after motile sperm isolation by swim-up. Fertility and Sterility, 60(6):1046-1051.

[9] JOHNSON, L.A. 1991. Sex preselection in swine: altered sex ratios in offspring following surgical insemination of flow sorted X- and Y-bearing sper. Reprod. Dom. Animal, 26:309-314.

[10] JOHNSON, L. A. 1992. Gender preselection in domestic animals using flow cytometrically sorted sperm. J. Anim. Sci, v.70, suppl.2, p. 8-18.

[11] KOO, G.C., STACHPOLE, C.W., BOYSE, E.A. et al. 1973. Topographical location of H-Y antigen on mouse spermatozoa by immunoelectronmicroscopy. Proc. Nat. Acad. Sci., 70:1502-1505.

[12] MEISTRICH, M. L. 1982. Potential and limitation of physical methods for separation of sperm bearing an X- or Y- chromosome. In: Prospects for sexing mammalian sperm (R.P. Amann and G. E. Seidel eds.), Colorado Associated University Press, Boulder, Colorado, p. 157-168.

[13] MORUZZI, J.F. 1979. Selecting a mammalian species for the separation of X- and Y-chromosome-bearing spermatozoa. J. Reprod. Fertil., 37:319-323.

[14] PETER, A. J., JONES, P.P., ROBINSON, J.P. 1993. Fraction of bovine spermatozoa for sex selection: A rapid immunomagnetic technique to remove spermatozoa that contain the H-Y antigen. Theriogenology, 40:1177-1185.

[15] UTSUMI, K., HAYASHI, M., TAKAKURA, R. et al. 1993. Embryo sex selection by a rat male-specific antibody and the cytogenetic and developmental confirmation in cattle embryos. Mol. Reprod. and Devel., 34:25-32.

[16] VAN VLECK, L.D., EVERETT, R. W. 1975. Genetic value of sexed semen to produce dairy heifers. J. Dairy Sci., 59(10):1802-1808.

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Anticorpo monoclonal contra uma proteína macho-específica de 19 kDa em espermatozóides bovinos: uma metodologia promissora para imunosexagem

Monoclonal antibody against male-specific protein of 19 KDa from bovine spermatozoa: a successful methodology for imunosexing

Resumos

Datas de Publicação

Histórico