A monoclonal antibody against human kidney gamma-glutamyl transpeptidase: preparation, immunochemical, and immunohistochemical characterization.

Abstract

To perform immunohistochemical study of the distribution of gamma-glutamyl transpeptidase in human organs, a highly specific antibody against the human enzyme is required. We prepared monoclonal antibody against gamma-glutamyl transpeptidase from human kidney, using the hybridoma technique. The antibody was of the IgG1 type and the light chain belonged to the kappa class. The antibody reacted specifically with the 63 KD heavy subunit of the enzyme. Examination of the specificity of the antibody performed by immunohistochemical staining of human kidney sections revealed that the antigen was localized on the brush border and along the basolateral membrane of the epithelial cells of both the convoluted and the straight portions of the proximal tubule. This antibody was also reactive in several human organs other than kidney, including epididymis, prostate, seminal vesicle, pancreas, and normal liver, and in human hepatoma. These findings indicate the existence of an antigenic determinant common to human kidney and other organs. The monoclonal antibody did not crossreact with mouse, rat, guinea pig, rabbit, or pig kidney.

Immunohistochemical detection of gamma-glutamyl transpeptidase in normal human tissue.

“…Detection of high levels of GGT in the proximal tubules of the kidney by the GGT129 antibody is in agreement with previous studies of GGT expression in humans and rodents (Shiozawa et al, 1989;Rutenburg et al, 1969;Albert et al, 1964). In the kidney, GGT initiates the cleavage of glutathione into three amino acids which can then be reabsorbed from the glomerular filtrate.…”

“…A Western blot of homogenates of five human tissues showed that the affinity-purified antibody reacted with a single band corresponding to the large subunit of GGT (Shiozawa et al, 1989;Bte and Galbraith, 1987). The amount of GGT heavy subunit detected on the Western blot correlated with the relative level of GGT activity determined biochemically and by immunohistochemical staining of the corresponding tissues.…”

“…Many human tissues have never been analyzed for GGT expression. Among tissues that have been analyzed there are conflicting results regarding the localization and level of GGT expression (Shiozawa et al, 1989;Albert et al, 1964;Glenner et al. 1962;Goldbarg et al, 1960).…”

Monoclonal Antibodies against Rat Kidney Antigens

“…aminopeptidases A and M or angiotensinase A and dipeptidyl peptidase lv. which form a major group of cell-membrane-bound proteins in the renal tubular brush border [11,14].…”

Monoclonal antibody to a common antigen of secretory granule membranes: intracellular localization and recycling of the antigen after secretion.

“…The method of MAb production has been described previously (38,45). In short, 6-week-old female Balb/c mice were subcutaneously immunized with the secretory granule membrane (100 pg protein) with complete Freund's adjuvant, and with incomplete Freund's adjuvant 2 weeks later.…”