Stability of Hormone Receptors with Fixation: Implications for Immunocytochemical Localization of Receptors*

Salih, H.; Murthy, Ganti S.; Friesen, Henry G.

doi:10.1210/endo-105-1-21

Cited by 7 publications

(4 citation statements)

References 11 publications

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“…The use of fresh, frozen tissue with only a brief fixation of sections in acetone has been found to be essential. This is in accordance with the findings of Salih et at. (1979) who assessed the binding of ovine prolactin to tissues after pretreatment with a range of fixatives and noted a reduction in specific prolactin binding with an increase in non-specific binding after fixation.…”

Section: Discussionsupporting

confidence: 94%

“…The purity of the human prolactin, the specificity of the antiserum to human prolactin and the absence of staining in sections treated with absorbed specific antiserum or non-immune serum are all good evidence that prolactin binding sites have been demonstrated in breast tissue in this study. Salih et al (1979) have suggested two further controls to assess specificity in immunohistochemical studies. An antiserum to prolactin receptor can be used to block the binding site.…”

Section: Discussionmentioning

confidence: 99%

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The localization of prolactin binding sites in human breast tissue

Dhadly¹,

Walker²

1983

Intl Journal of Cancer

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An immunohistochemical method involving the application of purified human prolactin and a specific antiserum to human prolactin, followed by the peroxidase-anti-peroxidase immunoperoxidase technique, has been used to detect prolactin binding in benign and malignant human breast tissue. The use of fresh, frozen material has been found to be essential. Prolactin binding has been shown to be a consistent feature of benign breast tissue but a variation within samples has been noted which is irrespective of hyperplastic changes. Fifty six percent of breast carcinomas gave a positive reaction but heterogeneity of binding has been shown to be a significant feature. A relationship between the presence and extent of prolactin binding and good histological differentiation of the tumours has been noted. It is concluded that immunohistochemistry is a suitable method for the demonstration of prolactin binding sites in human breast tissue and can provide useful information with regard to tumour heterogeneity.

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Section: Discussionsupporting

confidence: 94%

Section: Discussionmentioning

confidence: 99%

The localization of prolactin binding sites in human breast tissue

Dhadly¹,

Walker²

1983

Intl Journal of Cancer

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“…Neither, however, reported whether the binding characteristics were altered by 4% PAF. Salih et al (1979) looked a t the effect of several fixatives, used for immunocytochemistry, on binding to prolactin receptors in liver homogenates. They found that fixation with either Lillies' buffered formalin or 0.02% picric acid plus 2% formaldehyde did not significantly change specific binding, nonspecific binding, or K,, although both decreased B, , , by about 40%.…”

Section: Discussionmentioning

confidence: 99%

Effect of paraformaldehyde fixation on localization and characterization of insulin‐like growth factor‐I (IGF‐I) receptors in the rat brain

King

Baskin

1991

Anat. Rec.

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In order to design an approach for localizing IGF-I receptors within the intact CNS, the effect of paraformaldehyde (PAF) fixation on receptor binding was examined. Cryostat sections of rat brains, which were perfused with 0 to 10% PAF, were incubated in 125I-IGF-I and assayed for binding under equilibrium conditions. Binding was quantified from 10 brain regions, involving laminae and nuclei from median eminence, thalamus, hippocampus, choroid plexus, pyriform cortex, and cerebral cortex, by computer densitometry of film autoradiographs. The specific binding, saturation curves, Bmax and Ka, ligand specificity, and binding reversibility of IGF-I binding sites were not significantly affected by 1% or 2% PAF. However, 4% PAF elevated IGF-I receptor total binding, nonspecific binding, and Ka, and decreased Bmax, presumably by increasing the number of tissue-receptor interconnections. Only nonspecific 125I-IGF-I binding persisted when 10% PAF was used. These results indicate that tissue perfused with 2% PAF can be used for localizing IGF-I receptors by autoradiographic binding methods.

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“…This is reminiscent ofthe unique ability ofpicric acid/ paraformaldehyde fixatives to preserve the binding activity of peptide hormone receptors (2,3,7). Furthermore, even in the amine reagent-treated immunoglobulin molecule, the hypervariable regions seemed to be more resistant to destruction of their ability for reaction with antigen than are epitopes in other molecular regions for reaction with second antibody (8,9).…”

Section: Discussionmentioning

confidence: 99%

A new hypothalamic substance, and not luteinizing hormone-releasing hormone, is detected immunocytochemically by antibody to luteinizing hormone-releasing hormone.

Sternberger¹,

Greenwald²,

Hock³

et al. 1981

Proc. Natl. Acad. Sci. U.S.A.

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A new hypothalamic substance, and not luteinizing hormonereleasing hormone, is detected immunocytochemically by antibody to luteinizing hormone-releasing hormone ( Scharrer, April 13, 1981 ABSTRACT Adjacent paraffin sections of rat hypothalami fixed in Bouin's fluid were treated either with buffer or with luteinizing hormone-releasing hormone (LHRH) before immunocytochemical staining with anti-LHRH. Upon buffer pretreatment, pituitary gonadotrophs were unstained and hypothalamic fibers were stained. Upon LHRH pretreatment, pituitary gonadotrophs were stained (receptor reaction) and hypothalamic fibers were unstained. Extension of washes and use of series of neutralizing antisera between LHRH application and immunocytochemical staining, as well as the absence of inhibiting concentrations of LHRH in the later washes and neutralizing antisera removed from the sections, excluded the possibility that the disappearance of visualization of hypothalamic fibers was due to blockage of anti-LHRH in immunocytochemical staining. The results suggested that LHRH removed from the sections an immunocytochemically stainable but as yet unknown analog of LHRH and replaced it with LHRH, which in turn became lost during subsequent immunocytochemical processing. This idea was confirmed by the isolation by high-pressure liquid chromatography of a peak, distinct from LHRH, upon treatment of hypothalami with LHRH. It is suggested that the new substance may be carrierheld and that this substance, rather than LHRH, is normally detected by immunocytochemistry with anti-LHRH. Added LHRH binds not only to high-affinity pituitary receptors but also to lowaffinity hypothalamic carriers. Projections presumed to contain the decapeptide luteinizing hormone-releasing hormone (LHRH) are revealed by immunocytochemistry with anti-LHRH over wide areas in the brain (for review, see ref. 1). Visualization ofLHRH in Vibratome and even paraffin sections of fixed tissue is surprising because the decapeptide possesses no primary amine that is readily reactive with common aldehyde fixatives. Also, LHRH is highly soluble in water and in the alcohols used during fixation, embedding, and immunocytochemical staining.

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Stability of Hormone Receptors with Fixation: Implications for Immunocytochemical Localization of Receptors*

Cited by 7 publications

References 11 publications

The localization of prolactin binding sites in human breast tissue

The localization of prolactin binding sites in human breast tissue

Effect of paraformaldehyde fixation on localization and characterization of insulin‐like growth factor‐I (IGF‐I) receptors in the rat brain

A new hypothalamic substance, and not luteinizing hormone-releasing hormone, is detected immunocytochemically by antibody to luteinizing hormone-releasing hormone.

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