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Titolo:
Immunolocalization of the electrogenic Na+-HCO3- cotransporter in mammalian and amphibian kidney
Autore:
Schmitt, BM; Biemesderfer, D; Romero, MF; Boulpaep, EL; Boron, WF;
Indirizzi:
Yale Univ, Sch Med, Dept Cellular & Mol Physiol, New Haven, CT 06520 USA Yale Univ New Haven CT USA 06520 r & Mol Physiol, New Haven, CT 06520 USA YaleAUniv, Sch Med, Dept Internal Med, Nephrol Sect, New Haven, CT 06520 US Yale Univ New Haven CT USA 06520 ed, Nephrol Sect, New Haven, CT 06520 US
Titolo Testata:
AMERICAN JOURNAL OF PHYSIOLOGY-RENAL PHYSIOLOGY
fascicolo: 1, volume: 45, anno: 1999,
pagine: F27 - F38
SICI:
0363-6127(199901)45:1<F27:IOTENC>2.0.ZU;2-9
Fonte:
ISI
Lingua:
ENG
Soggetto:
RENAL PROXIMAL TUBULE; THICK ASCENDING LIMB; FUNCTIONAL EXPRESSION; BICARBONATE TRANSPORT; DISTAL TUBULE; PH REGULATION; RAT-KIDNEY; SODIUM; CELLS; ACIDIFICATION;
Keywords:
fluid and electrolytes; renal bicarbonate reabsorption; acid-base; polyclonal antibody; immunofluorescence; immunoblotting; rat; rabbit; salamander;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
36
Recensione:
Indirizzi per estratti:
Indirizzo: Schmitt, BM YaleTUniv, Sch Med, Dept Cellular & Mol Physiol, 333 Cedar St,New Haven, C Yale Univ 333 Cedar St New Haven CT USA 06520 St, New Haven, C
Citazione:
B.M. Schmitt et al., "Immunolocalization of the electrogenic Na+-HCO3- cotransporter in mammalian and amphibian kidney", AM J P-REN, 45(1), 1999, pp. F27-F38

Abstract

Electrogenic cotransport of Na+ and HCO3- is a crucial element of HCO3- reabsorption in the renal proximal tubule (PT). An electrogenic Na+-HCO3- cotransporter (NBC) has recently been cloned from salamander and rat kidney. In the present study, we generated polyclonal antibodies (pAbs) to NBC and used them to characterize NBC on the protein level by immunochemical methods. We generated pAbs in guinea pigs and rabbits by immunizing with a fusion protein containing the carboxy-terminal 108 amino acids (amino acids 928-1035) of rat kidney NBC (rkNBC). By indirect immunofluorescence microscopy, the pAbs strongly labeled HEK-293 cells transiently expressing NBC, but not in untransfected cells. By immunoblotting, the pAbs recognized a similar to130-kDa band in Xenopus laevis oocytes expressing rkNBC, but not in control oocytes injected with water or cRNA for the Cl-/HCO3- exchanger AE2. In immunoblotting experiments on renal microsomes, the pAbs specifically labeled a major band at similar to 130 kDa in both rat and rabbit, as well as a single similar to 160-kDa band in salamander kidney. By indirect immunofluorescence microscopy on 0.5-mu m cryosections of rat and rabbit kidneys fixedin paraformaldehyde-lysine-periodate (PLP), the pAbs produced a strong andexclusively basolateral staining of the PT. In the salamander kidney, the pAbs labeled only weakly the basolateral membrane of the PT. In contrast, we observed strong basolateral labeling in the late distal tubule, but not in the early distal tubule. The specificity of the pAbs for both immunoblotting and immunohistochemistry was confirmed in antibody preabsorption experiments using either the fusion protein used for immunization or similarly prepared control fusion proteins. In summary, we have developed antibodies specific for NBC, determined the apparent molecular weights of rat, rabbit, and salamander kidney NBC proteins, and described the localization of NBC within the kidney of these mammalian and amphibian species.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 01/10/20 alle ore 13:33:51