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Characterization and use of polyclonal antibody to Na + , K + ‐ATPase: Immunocytochemical localization in salt glands of the duck
Author(s) -
Russo James J.,
Merchant Juanita L.,
Eager Patricia R.,
Barrnett Russell J.
Publication year - 1987
Publication title -
cell biochemistry and function
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.933
H-Index - 61
eISSN - 1099-0844
pISSN - 0263-6484
DOI - 10.1002/cbf.290050102
Subject(s) - polyclonal antibodies , microbiology and biotechnology , antibody , immunocytochemistry , salt (chemistry) , salt gland , chemistry , atpase , biology , biochemistry , enzyme , immunology , endocrinology , secretion
Abstract The amount of Na + , K + ‐ATPase of the avian salt gland increased concomitantly with plasma membrane surface area during salt feeding of ducklings (adaptation), and both enzyme content and membrane surface area decreased upon return to fresh water (deadaptation). In a further study of the enzyme, a marker for plasma membrane biogenesis, polyvalent antibodies were raised to the denatured α‐subunit of the purified ATPase. Antisera did not inhibit enzymatic activity but immunoprecipitated the phosphorylated intermediate of the α‐subunit. Furthermore, the α‐subunit, which was not glycosylated, was immunoprecipitated from homogenates of tissue slices metabolically labelled with [ 35 S]‐methionine, using antisera raised against either duck salt gland or dog kidney α‐subunit. The former antisera also recognized the α‐subunit in the brain, heart, kidney, liver, intestine and skeletal muscle of the duck. Immunocytochemistry with the antisera raised to the duck salt gland α‐subunit revealed reaction at basolateral as well as apical plasma membrane in the duck salt gland principal cells, with essentially no deposits no deposits on peripheral cells, fibroblasts, erythrocytes, endothelial cells and neural elements. Within the principal cells, immunolabelling was also detected on small vesicles, multivesicular bodies and lysosomes; deposits on extracellular debris and vesicles in the lateral and lumenal spaces were also apparent. The labelling patterns were qualitatively but not quantitatively similar in salt glands of control, adapted and deadapted ducklings, and are discussed in the context of a model for plasma membrane biogenesis and turnover in which degradative events may play a major role.