Métodos de coloração de Roeser (1972): modificado - e Kropp (1972) visando a substituição do azul de astra por azul de alcião 8GS ou 8GX

Luque, Rebeca; Sousa, Hildeberto C. de; Kraus, Jane Elizabeth

doi:10.1590/S0102-33061996000200001

Resumos

Em folhas de Lychnophora ericoides, Coffea arabica e Nymphaea mexicana foram testados métodos de coloração baseados em Roeser (1972) modificado e Kropp (1972), visando a substituição do corante azul de astra por azul de alcião 8GS ou 8GX. As amostras foram fixadas em FAA, desidratadas em série butílica terciária e incluídas em parafina. Os cortes histológicos transversais foram corados segundo diferentes baterias de coloração, modificadas quanto ao tipo de corante usado, diferenciador e série de desidratação. As lâminas permanentes foram preparadas com bálsamo-do-canadá sintético. Os resultados obtidos evidenciaram que o método de Roeser (1972) modificado é melhor que o de Kropp (1972), nas condições deste experimento. O azul de astra pode ser substituído por azul de alcião 8GX e a desidratação pode ser em série isopropílica ou etílica, sem grande diferença entre elas. São discutidos os resultados provenientes das diferentes colorações.

método de coloração; azul de astra; azul de alcião; fucsina básica; safranina; histologia vegetal

Staining methods based on modified Roeser (1972) as well as that of Kropp (1972) were done with leaves of Lychnophora ericoides, Coffea arabica and Nymphaea mexicana aiming at substituing the astra blue stain by alcian blue 8GS or 8GX. Samples were fixed in FAA, dehydrated in teit-butyl alcohol series and embedded in paraffin. The histological transversal sections were stained in different staining batteries, modified according to the type of stains and differentiators, and the series of dehydration. Permanent slides were prepared with synthetic Canada balsam. The results obtained indicate that the modified Roeser method (1972) is better than that of Kropp (1972) under present experimental conditions. The astra blue can be substituted by alcian blue 8GX and dehydration can be isopropylic or ethylic, with little difference between them. The results obtained with the different staining techniques used are discussed.

staining method; astra blue; alcian blue; basic fuchsin; safranin; plant histology

Métodos de coloração de Roeser (1972) - modificado - e Kropp (1972) visando a substituição do azul de astra por azul de alcião 8GS ou 8GX

Staining methods of modified Roeser (1972) and Kropp (1972), aiming at substituing the astra blue by alcian blue 8GS or 8GX

Rebeca Luque^I; Hildeberto C. de Sousa^II; Jane Elizabeth Kraus^III

^IDepart. de Biologia. Facultad de Ciências, Universidad de Los Andes, La Hechicera. Mérida 5101, Venezuela

^IIDepart. de Ciências Biológicas. Instituto de Ciências Exatas e Biológicas, Universidade Federal de Ouro Preto, Campus do Morro do Cruzeiro, CEP 35400-000 Ouro Preto, MG. Brasil

^IIIDepart. de Botânica, Instituto de Biociências, Universidade de São Paulo, C.P. 11461, CEP 05422-970 São Paulo, SP. Brasil

RESUMO

Em folhas de Lychnophora ericoides, Coffea arabica e Nymphaea mexicana foram testados métodos de coloração baseados em Roeser (1972) modificado e Kropp (1972), visando a substituição do corante azul de astra por azul de alcião 8GS ou 8GX. As amostras foram fixadas em FAA, desidratadas em série butílica terciária e incluídas em parafina. Os cortes histológicos transversais foram corados segundo diferentes baterias de coloração, modificadas quanto ao tipo de corante usado, diferenciador e série de desidratação. As lâminas permanentes foram preparadas com bálsamo-do-canadá sintético. Os resultados obtidos evidenciaram que o método de Roeser (1972) modificado é melhor que o de Kropp (1972), nas condições deste experimento. O azul de astra pode ser substituído por azul de alcião 8GX e a desidratação pode ser em série isopropílica ou etílica, sem grande diferença entre elas. São discutidos os resultados provenientes das diferentes colorações.

Palavras-chave: método de coloração, azul de astra. azul de alcião, fucsina básica, safranina, histologia vegetal

ABSTRACT

Staining methods based on modified Roeser (1972) as well as that of Kropp (1972) were done with leaves of Lychnophora ericoides, Coffea arabica and Nymphaea mexicana aiming at substituing the astra blue stain by alcian blue 8GS or 8GX. Samples were fixed in FAA, dehydrated in teit-butyl alcohol series and embedded in paraffin. The histological transversal sections were stained in different staining batteries, modified according to the type of stains and differentiators, and the series of dehydration. Permanent slides were prepared with synthetic Canada balsam. The results obtained indicate that the modified Roeser method (1972) is better than that of Kropp (1972) under present experimental conditions. The astra blue can be substituted by alcian blue 8GX and dehydration can be isopropylic or ethylic, with little difference between them. The results obtained with the different staining techniques used are discussed.

Key words: staining method, astra blue, alcian blue, basic fuchsin, safranin, plant histology

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Agradecimentos

Ao CNPq, pela bolsa de pesquisa concedida (Proc. nº 301776/83). Aos Professores Doutores Berta Lange de Morretes, Liliana Forneris e Paulo Alberto Otto, pela tradução dos textos em alemão.

Recebido em 14/12/95.

Aceito em 2/6/96.

Referências bibliográficas

Achar, B.N.; Bhandari, J.M. & Urs, H.G.V.G. 1993. A rapid safranin-metal phthalocyanine double staining technique for plants. Biotech. & Histochem. 68: 127-131.
Adams, C.W.M. & Sloper, J.C. 1956. The hypothalamic elaboration of posterior pituitary principles in man, the rat and dog; histochemical evidence derived from a performic acid-alcian blue reaction for cystine. J. Endocr. 13: 221-228.
Bene, K. 1968. On the stainability of plant cell walls with alcian blue. Biol. Plant. 10: 334-346.
Berlyn, G.P. & Miksche, J.P. 1976. Botanical microtechnique and cytochemistry. Ames: Iowa State University.
Braga, M.M.N. 1977. Anatomia foliar de Bromeliaceae da Campina. Acta Amazonica 7: 1-74, suplemento.
Braga, M.M.N. 1982. Aspectos anatômicos das folhas de Virola Aubl. (Myristicaceae) como contribuição para sua Taxonomia. Manaus (AM): Instituto Nacional de Pesquisas da Amazônia e Fundação Universidade da Amazônia. Tese de Doutorado.
Bukatsch, F. 1972. Bemerkungen zur Doppelfärbung: Astrablau-Safranin. Mikrokosmos 61: 255.
Chaubal, R.; Wilmot, V.A. & Wynn, W.K. 1991. Visualization, adhesiveness, and cytochemistry of the extracellular matrix produced by urediniospore germ tubes of Puccinia sorghi. Can. J. Bot. 69: 2044-2054.
Clark, G. 1981. Staining procedures. 4Ş ed. Baltimore: Williams & Wilkins.
Decker, B. & Reale, E. 1991. The glomerular filtration barrier of the kidney in seven vertebrates classes: comparative morphological and histochemical observations. Eur. J. Basic and Appl. Histochem. 35: 15-36.
Gahan, P.B. 1984. Plant histochemistry and cytochemistry: an introduction. London: Academic Press.
Gerlach, D. 1984. Botanische Mikrotechnik: eine Einführung. Sttutgart: Georg Thieme Verlag.
Gilloteaux, J.; Pomerants, B.J., Kelly, T.R., Menu, R., Pelletier, G. & Vanderhaeghen, J. J. 1990. Light and eletron microscopical immunolocalization of neuropeptide Y-containing nerves in the hamster gallbladder. Biol. Struct. Morphog. 3: 89-96.
Horobin, R.W. & Goldstein, D.J. 1972. Impurities and staining characteristics of alcian blue samples. Histochem. J. 4: 391-399.
Jeffree, C.E. 1993. Immobilization of pectin, xyloglucan and other soluble plant polysaccharides on blotting membranes. New Phytol. 125: 695-706.
Jenkins, P.G.; Pulsford, A.L. & Harris, J.E. 1992. Microscopy of the absorptive cells and gut-associated lymphoid tissue of the flounder Platichthys flesus. J. Mar. Biol. Assoc. U.K. 72: 553-567.
Johansen, D.A. 1940. Botanical microtechnique. New York: McGraw-Hill Book.
Kropp, U. 1972. Leitbündel. Mikrokosmos 61: 342-345.
Lillie, R.D. 1990. H. J. Conn's biological stains. 9.Ş ed. St. Louis: Sigma Chemical.
Lindauer, R. 1971. Die Pflanzen-Epidermis: Isolierung, Färbung, Einschluss. Mikrokosmos 60: 170-178.
Lison, L. 1954. Alcian blue 8G with chlorantine fast red 5B. A technic for selective staining of mucopolysaccharides. Stain Technol. 29: 131-138.
Maácz, G.J. & Vágás, E. 1961. A new method for staining of cellulose and lignified cells-walls. Mikroskopie 16: 40-43.
Malzacher, P. 1972. Eine neue Färbung für zoologische Totalpräparate: Astrablau-Boraxkarmin. Mikrokosmos 61: 181-182.
McManus, J.F.A. & Mowry, R.W. 1960. Staining methods histological and histochemical. New York: Harper & Row.
Mowry, R.W. 1956. Alcian blue technics for the histochemical study of acidic carbohydrates. Proc. Histochem. Soc. 4: 407.
Mowry, R.W. 1960. Revised method producing improved coloration of acidic polyssacharides with alcian blue 8GX supplied currently. Proc. Histochem. Soc. 8: 323-324.
Niimura, S.; Fujisawa, N.; Sato, N.L. & Ishida, K. 1991. Histochemical observations of proteins, polysaccharides and lipids in the oocytes and embryos of mastomys (Praomys natalensis) during the folliculogenesis and early development. Japan. J. Animal Reprod. 37: 65-70.
Pickett-Heaps, J.; Hill, D.R.A. & Blaze, K.L. 1991. Active gliding motility in an araphid marine diatom, Ardissonea (formerly Synedra) crystallina. J. Phycol. 27: 718-725.
Pioch, W. 1957. Über die Darstellung saurer Mucopolysaccharide mit dem Kupferphthalocyaninfarbstoff Astrablau. Virchows Arch. 330: 337-346.
Roeser, K.R. 1972. Die Nadel der Schwarzkiefer - Massenproduckt und Kunstwerk der Natur. Mikrokosmos 61: 33-36.
Rubio, C.A.; Huang, C.B.; Porwit, A., Kock., Y. & Stenling, R. 1991. Quantification of the colonic mucous cell population during protracted stress in rats. Stress Med. 7: 145-152.
Schumacher, U. & Adam, E. 1994. Standardization of staining in glycosaminoglycan histochemistry: alcian blue, its analogues and diamine methods. Biotech. & Histochem. 69: 18-24.
Serrato-Valenti, G.; Cornara, L., Lotito, S. & Quagliotti, L. 1992. Seed coat structure and histochemistry of Abelmoschus esculentus: chalazal region and water entry. Ann. Bot. (Lond.) 69: 313-321.
Steesman, H.F. 1950. Alcian blue 8GS: a new stain for mucin. Q.J. Microsc. Sei. 91: 477-479.
Srebotnik, E. & Messner, K. 1994. A simple method that uses differential staining and light microscopy to assess the selectivity of wood delignification by white rot fungi. App. Environ. Microbiol. 60: 1383-1386.
Tanigawa, K.; Kawaguchi, M., Tanaka, O. & Kato, Y. 1991. Skeletal malformations in rat offspring: long-term effect of maternal insulin-induced hypoglycemia during organogenesis. Diabetes 40: 1115-1121.
Tas, J. 1977a. The alcian blue and combined alcian blue-safranin O staining of glycosaminoglicans studied in a model system and in mast cells. Histochem. J. 9: 205-230.
Tas, J. 1977b. Polyacrilamide films as a tool for investigating qualitative and quantitative aspects of the staining of glycosaminoglycans with basic dyes. Histochem. J. 9: 267-276.
Wismar, B.I. 1966. Quad-type stain for the simultaneous demonstration of intracellular and extracellular tissue components. Stain Technol. 41: 309-313.
Woessner, E. 1970. Astrablau-Safranin - noch einfacher. Botanische Mikropräparate mit geringstem Aufwand. Mikrokosmos 59: 31-32.

Datas de Publicação

Publicação nesta coleção
14 Out 2011
Data do Fascículo
Dez 1996

Histórico

Recebido
14 Dez 1995
Aceito
02 Jun 1996

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

[1] Achar, B.N.; Bhandari, J.M. & Urs, H.G.V.G. 1993. A rapid safranin-metal phthalocyanine double staining technique for plants. Biotech. & Histochem. 68: 127-131.

[2] Adams, C.W.M. & Sloper, J.C. 1956. The hypothalamic elaboration of posterior pituitary principles in man, the rat and dog; histochemical evidence derived from a performic acid-alcian blue reaction for cystine. J. Endocr. 13: 221-228.

[3] Bene, K. 1968. On the stainability of plant cell walls with alcian blue. Biol. Plant. 10: 334-346.

[4] Berlyn, G.P. & Miksche, J.P. 1976. Botanical microtechnique and cytochemistry. Ames: Iowa State University.

[5] Braga, M.M.N. 1977. Anatomia foliar de Bromeliaceae da Campina. Acta Amazonica 7: 1-74, suplemento.

[6] Braga, M.M.N. 1982. Aspectos anatômicos das folhas de Virola Aubl. (Myristicaceae) como contribuição para sua Taxonomia. Manaus (AM): Instituto Nacional de Pesquisas da Amazônia e Fundação Universidade da Amazônia. Tese de Doutorado.

[7] Bukatsch, F. 1972. Bemerkungen zur Doppelfärbung: Astrablau-Safranin. Mikrokosmos 61: 255.

[8] Chaubal, R.; Wilmot, V.A. & Wynn, W.K. 1991. Visualization, adhesiveness, and cytochemistry of the extracellular matrix produced by urediniospore germ tubes of Puccinia sorghi. Can. J. Bot. 69: 2044-2054.

[9] Clark, G. 1981. Staining procedures. 4Ş ed. Baltimore: Williams & Wilkins.

[10] Decker, B. & Reale, E. 1991. The glomerular filtration barrier of the kidney in seven vertebrates classes: comparative morphological and histochemical observations. Eur. J. Basic and Appl. Histochem. 35: 15-36.

[11] Gahan, P.B. 1984. Plant histochemistry and cytochemistry: an introduction. London: Academic Press.

[12] Gerlach, D. 1984. Botanische Mikrotechnik: eine Einführung. Sttutgart: Georg Thieme Verlag.

[13] Gilloteaux, J.; Pomerants, B.J., Kelly, T.R., Menu, R., Pelletier, G. & Vanderhaeghen, J. J. 1990. Light and eletron microscopical immunolocalization of neuropeptide Y-containing nerves in the hamster gallbladder. Biol. Struct. Morphog. 3: 89-96.

[14] Horobin, R.W. & Goldstein, D.J. 1972. Impurities and staining characteristics of alcian blue samples. Histochem. J. 4: 391-399.

[15] Jeffree, C.E. 1993. Immobilization of pectin, xyloglucan and other soluble plant polysaccharides on blotting membranes. New Phytol. 125: 695-706.

[16] Jenkins, P.G.; Pulsford, A.L. & Harris, J.E. 1992. Microscopy of the absorptive cells and gut-associated lymphoid tissue of the flounder Platichthys flesus. J. Mar. Biol. Assoc. U.K. 72: 553-567.

[17] Johansen, D.A. 1940. Botanical microtechnique. New York: McGraw-Hill Book.

[18] Kropp, U. 1972. Leitbündel. Mikrokosmos 61: 342-345.

[19] Lillie, R.D. 1990. H. J. Conn's biological stains. 9.Ş ed. St. Louis: Sigma Chemical.

[20] Lindauer, R. 1971. Die Pflanzen-Epidermis: Isolierung, Färbung, Einschluss. Mikrokosmos 60: 170-178.

[21] Lison, L. 1954. Alcian blue 8G with chlorantine fast red 5B. A technic for selective staining of mucopolysaccharides. Stain Technol. 29: 131-138.

[22] Maácz, G.J. & Vágás, E. 1961. A new method for staining of cellulose and lignified cells-walls. Mikroskopie 16: 40-43.

[23] Malzacher, P. 1972. Eine neue Färbung für zoologische Totalpräparate: Astrablau-Boraxkarmin. Mikrokosmos 61: 181-182.

[24] McManus, J.F.A. & Mowry, R.W. 1960. Staining methods histological and histochemical. New York: Harper & Row.

[25] Mowry, R.W. 1956. Alcian blue technics for the histochemical study of acidic carbohydrates. Proc. Histochem. Soc. 4: 407.

[26] Mowry, R.W. 1960. Revised method producing improved coloration of acidic polyssacharides with alcian blue 8GX supplied currently. Proc. Histochem. Soc. 8: 323-324.

[27] Niimura, S.; Fujisawa, N.; Sato, N.L. & Ishida, K. 1991. Histochemical observations of proteins, polysaccharides and lipids in the oocytes and embryos of mastomys (Praomys natalensis) during the folliculogenesis and early development. Japan. J. Animal Reprod. 37: 65-70.

[28] Pickett-Heaps, J.; Hill, D.R.A. & Blaze, K.L. 1991. Active gliding motility in an araphid marine diatom, Ardissonea (formerly Synedra) crystallina. J. Phycol. 27: 718-725.

[29] Pioch, W. 1957. Über die Darstellung saurer Mucopolysaccharide mit dem Kupferphthalocyaninfarbstoff Astrablau. Virchows Arch. 330: 337-346.

[30] Roeser, K.R. 1972. Die Nadel der Schwarzkiefer - Massenproduckt und Kunstwerk der Natur. Mikrokosmos 61: 33-36.

[31] Rubio, C.A.; Huang, C.B.; Porwit, A., Kock., Y. & Stenling, R. 1991. Quantification of the colonic mucous cell population during protracted stress in rats. Stress Med. 7: 145-152.

[32] Schumacher, U. & Adam, E. 1994. Standardization of staining in glycosaminoglycan histochemistry: alcian blue, its analogues and diamine methods. Biotech. & Histochem. 69: 18-24.

[33] Serrato-Valenti, G.; Cornara, L., Lotito, S. & Quagliotti, L. 1992. Seed coat structure and histochemistry of Abelmoschus esculentus: chalazal region and water entry. Ann. Bot. (Lond.) 69: 313-321.

[34] Steesman, H.F. 1950. Alcian blue 8GS: a new stain for mucin. Q.J. Microsc. Sei. 91: 477-479.

[35] Srebotnik, E. & Messner, K. 1994. A simple method that uses differential staining and light microscopy to assess the selectivity of wood delignification by white rot fungi. App. Environ. Microbiol. 60: 1383-1386.

[36] Tanigawa, K.; Kawaguchi, M., Tanaka, O. & Kato, Y. 1991. Skeletal malformations in rat offspring: long-term effect of maternal insulin-induced hypoglycemia during organogenesis. Diabetes 40: 1115-1121.

[37] Tas, J. 1977a. The alcian blue and combined alcian blue-safranin O staining of glycosaminoglicans studied in a model system and in mast cells. Histochem. J. 9: 205-230.

[38] Tas, J. 1977b. Polyacrilamide films as a tool for investigating qualitative and quantitative aspects of the staining of glycosaminoglycans with basic dyes. Histochem. J. 9: 267-276.

[39] Wismar, B.I. 1966. Quad-type stain for the simultaneous demonstration of intracellular and extracellular tissue components. Stain Technol. 41: 309-313.

[40] Woessner, E. 1970. Astrablau-Safranin - noch einfacher. Botanische Mikropräparate mit geringstem Aufwand. Mikrokosmos 59: 31-32.