Localization of Adrenocorticotropic Hormone by Histochemical and Immunochemical Methods

Abstract

The fluorescent antibody technique was adapted to the localization of native protein antigens in cells and tissues. This method was applied specifically to the localization of adrenocorticotropic hormone in the pituitary gland. An antiserum to hog ACTH was produced in an adrenalectomized rabbit. The γ2-globulin fraction of the serum was conjugated with fluorescein. After purification, the fluorescent antibody solution stained selectively the cytoplasm of basophil cells of the hog pituitary. No cells of sheep or beef pituitary or of hog kidney were stained. A fluorescent globulin solution prepared from normal rabbit serum gave no selective staining in any of these tissues. Immunochemical tests showed that the fluorescent antibody gave a precipitin reaction with a highly active ACTH preparation of low molecular weight. The supernatant solution from this reaction showed a loss of hormone activity.

Studies on Antibody Production

“…Fluorescence microscopy was carried out using the equipment previously described (5), modified by the use of a darkfield condenser (16,17).…”

“…[7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22], the spleen showed extensive nonspecific staining. A few persisting antibody-containing cells could not be excluded.…”

The Fine Structure of the Human Pituitary Chromophobe Adenoma with Special Reference to the Classification of this Tumor

“…The cell type concerning the production of ACTH has long been a matter of special controversy (FARQUHAR, 1957;FINERTY and BRISENO-CASTREJON, 1949;MARSHALL, 1951;LEZNOFF et al, 1962;RENNELS, 1964;SIPERSTEIN and ALISON, 1965;SMELSER, 1944 1) The longest dimension 2) The shortest dimension * Maximum diameter of the granules as determined from sectioned material ** Approximate diameter of the granules as conjectured from hormone assays ample, in the luteotroph. There may be a much more difference in the granule size between man and animal.…”

Functional Classification of Cell Types of the Anterior Pituitary Gland Accomplished by Electron Microscopy

“…Immunohistological studies Fluorescent antibody method was applied to the anterior pituitary gland in search of ACTH-cell by light microscopy (MARSHALL 1951, LEZNOFF et al 1962, but more detailed studies with the electron microscope using such immunohistological methods are required. Instead of a fluorescent substance, ferritin or peroxidase is conjugated to the antibody against a certain pituitary hormone for the use in electron microscopy.…”