Immunohistochemical study of .GAMMA.-glutamyl transpeptidase with monoclonal antibodies. I Preparation and characteristics of monoclonal antibodies to .GAMMA.-glutamyl transpeptidase.

Yasuda, Kenjiro; Yamashita, Shuji; Aiso, Sadakazu; Shiozawa, Masahide; Komatsu, Toshiko

doi:10.1267/ahc.19.589

Cited by 29 publications

(16 citation statements)

References 41 publications

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“…Antigen retrieval or exposure procedures were examined for the immunohistochemistry of both ER isoforms [12,41]. However, when the fixed frozen sections were autoclaved or microwaved in buffers, ERa-immunostaining using MC-20 antibody was reduced and ERb-immunostaining with Y-19 was abolished.…”

Section: Resultsmentioning

confidence: 99%

Ontogeny of Estrogen Receptor (ER) .ALPHA. and .BETA. in the Female Reproductive Tissues of Mice Neonatally Exposed to Diethylstilbestrol.

Yamashita¹

2002

Acta Histochem. Cytochem

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Ontogenical expression of estrogen receptor (ER) isoforms, ERa and ERb, was examined in order to study the roles of the isoforms in the morphogenesis of neonatal diethylstilbestrol (DES) treatment in the female reproductive tissues using an immunohistochemical method. The intensity of ERa immunostaining gradually increased with aging in the epithelial cells of oviducts, uteri, cervices and vaginae in the vehicle-treated control animals. Neonatal DES-treatment up-regulated the expression of ERa in the epithelial cells in these tissues and down-regulated it in the stromal cells at least until 10 days of age including the period of DES-treatment. On day 21, both control and DES-mice showed almost similar ERa distribution in these tissues. There were no differences in the ERa distribution in the ovaries of two experimental groups. ERa was localized in the theca cells and some stromal cells located interfollicularly. Neonatal DES exposure did not influence the distribution pattern of ERb protein during the period of treatment and postnatal development. The nuclear ERb-immunoreaction was seen in the granulosa cells of ovaries but not in the cells of reproductive tracts. These results demonstrated that ERa is the critical ER isoform for the estrogen-dependent responses in the normal female genital tract, and that the presence of ERa in the epithelial cells during the period of DES exposure should play important roles in the the induction of abnormal characteristics in the genital tract of DES-mice.

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Section: Resultsmentioning

confidence: 99%

Ontogeny of Estrogen Receptor (ER) .ALPHA. and .BETA. in the Female Reproductive Tissues of Mice Neonatally Exposed to Diethylstilbestrol.

Yamashita¹

2002

Acta Histochem. Cytochem

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“…Therefore, No heating PH 6 pH 6-9 pH 6-9-6 pH 9 pH 9-6 pH 9-6-9 heat-induced antigen retrieval procedures are considered to facilitate the interaction between antigens and antibodies by cleaving the cross-links. In addition, heating should also expose epitopes that are covered with oligomeric subunits or are located on the inner portion of the antigens regardless of formaldehyde fixation (Yasuda et al 1986;Pfund et al 1996;Yamashita et al 1997). …”

Section: Discussionmentioning

confidence: 99%

“…Pfund et al (1996) also reported that a monoclonal antibody prepared against bovine somatotropin reacted only with the denatured antigen. These results suggest that epitopes located on the inner portions of antigen molecules or those hidden by other oligomeric components must be exposed through denaturation before they can react with antibodies, regardless of treatment of fixatives (Yasuda et al 1986;Yamashita et al 1989Yamashita et al ,1997Robinson and Vandre 2001).…”

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confidence: 96%

Mechanisms of Heat-induced Antigen Retrieval: Analyses In Vitro Employing SDS-PAGE and Immunohistochemistry

Yamashita

Okada

2005

J Histochem Cytochem.

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S U M M A R YIn this study, we examined the mechanism of heat-induced antigen retrieval using analytical procedures involving SDS-PAGE, Western blotting, and immunohistochemistry. Five proteins were treated with 4% formaldehyde in the presence or absence of 25 mM CaCl 2 , then heated under various conditions after removal of formaldehyde and analyzed on SDS-PAGE. Formaldehyde produced inter-and intramolecular cross-links in the proteins. Heating at high temperatures cleaved these cross-links at all pH ranges examined (pH 3.0, 6.0, 7.5, 9.0) and produced almost the same electrophoregrams as the native proteins. Proteins treated with formaldehyde containing CaCl 2 showed similar electrophoretic patterns, observed without heating or after heating at pH 6.0 and pH 9.0 in the presence or absence of 10 mM EDTA. Western blot analyses demonstrated that the soluble forms of ␤ -actin (monomer and oligomers) and fibronectin were present in extracts from deparaffinized mouse uterine sections autoclaved for 15 min but not in extracts from unheated specimens. Nine of ten antigens, independent of their isoelectric points, exhibited much stronger immunoreaction in the sections heated at pH 9.0 than in those heated at pH 6.0. The second heating at pH 6.0 significantly decreased the immunostaining of the antigens that had been boiled at pH 9.0, but the immunostaining was recovered after a third heating at pH 9.0. These results suggest that the main mechanism of heat-induced antigen retrieval is disruption of the cross-links and that pH is an essential factor for a proper refolding of epitopes.

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“…In the previous study on the monoclonal antibody to rat kidney y-GTP, the antibody was reported to recognize the 51-kilodalton protein of the heavy subunit of the antigen (43), while the monoclonal antibody to rabbit kidney y-GTP, prepared in this study, reacted with the 54.5-kilodalton protein. Even though both antibodies recognized the protein of the heavy subunit of y-GTP of each animal, each antibody reacted with a different determinant, although the difference in the estimated values of the molecular weight of the polypeptides of two antigens recognized by each monoclonal antibody, 51 and 54.5 kilodaltons respectively, was small.…”

Section: Cross-reactivity Analysismentioning

confidence: 80%

“…The procedure used in the preparation is almost the same as that described in the previous report (43) . It is summarized as follows : 1) a microvilli fraction was prepared from the renal cortex, and solubilized by treatment with papain (27) ; 2) after centrifugation, the supernatant was brought to 65% saturation and thereafter to 95 % saturation with solid ammonium sulfate; 3) the crude enzyme preparation was further purified by column chromatography with Sephadex G-150.…”

Section: Methodsmentioning

confidence: 99%

Preparation of monoclonal antibody to rabbit kidney gamma-glutamyl transpeptidase and its cross-reactivity analysis in immunohistochemistry.

Komatsu

Yamashita

Yasuda

1987

Acta Histochem. Cytochem

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Immunohistochemical study of .GAMMA.-glutamyl transpeptidase with monoclonal antibodies. I Preparation and characteristics of monoclonal antibodies to .GAMMA.-glutamyl transpeptidase.

Cited by 29 publications

References 41 publications

Ontogeny of Estrogen Receptor (ER) .ALPHA. and .BETA. in the Female Reproductive Tissues of Mice Neonatally Exposed to Diethylstilbestrol.

Ontogeny of Estrogen Receptor (ER) .ALPHA. and .BETA. in the Female Reproductive Tissues of Mice Neonatally Exposed to Diethylstilbestrol.

Mechanisms of Heat-induced Antigen Retrieval: Analyses In Vitro Employing SDS-PAGE and Immunohistochemistry

Preparation of monoclonal antibody to rabbit kidney gamma-glutamyl transpeptidase and its cross-reactivity analysis in immunohistochemistry.

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