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Titolo:
IMMUNODISSECTION OF THE HUMAN PROXIMAL NEPHRON - FLOW SORTING OF S1S2S3, S1S2 AND S3 PROXIMAL TUBULAR CELLS
Autore:
HELBERT MJF; DAUWE SEH; VANDERBIEST I; NOUWEN EJ; DEBROE ME;
Indirizzi:
UNIV ANTWERP HOSP,DEPT NEPHROL,WILRIJKSTR 10 B-2650 ANTWERP BELGIUM UNIV ANTWERP,DEPT NEPHROL HYPERTENS B-2020 ANTWERP BELGIUM
Titolo Testata:
Kidney international
fascicolo: 2, volume: 52, anno: 1997,
pagine: 414 - 428
SICI:
0085-2538(1997)52:2<414:IOTHPN>2.0.ZU;2-W
Fonte:
ISI
Lingua:
ENG
Soggetto:
PLACENTAL ALKALINE-PHOSPHATASE; HUMAN-KIDNEY; EPITHELIAL-CELLS; MONOCLONAL-ANTIBODIES; GLYCOSYL-PHOSPHATIDYLINOSITOL; EXTRINSIC SIGNALS; MEMBRANE; CULTURE; EXPRESSION; ANTIGEN;
Keywords:
FLOW SORTING OF PT CELLS; PROXIMAL NEPHRON; CELL CULTURE; FLUORESCENCE-ACTIVATED CELL SORTING; IMMUNODISSECTION;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
48
Recensione:
Indirizzi per estratti:
Citazione:
M.J.F. Helbert et al., "IMMUNODISSECTION OF THE HUMAN PROXIMAL NEPHRON - FLOW SORTING OF S1S2S3, S1S2 AND S3 PROXIMAL TUBULAR CELLS", Kidney international, 52(2), 1997, pp. 414-428

Abstract

We report on the use of several proximal tubular cell (PTC) surface markers and corresponding antibodies in fluorescence-activated cell sorting (FACS), and their ability to identify and flow sort cells of defined proximal tubular origin (S1S2S3) or of defined proximal subsegmental origin (S1S2 only/S3 only). We tested monoclonal/polyclonal antibodies directed against five different surface peptidases [leucine aminopeptidase (LAP), neutral endopeptidase 24.11 (NEP), dipeptidyl peptidase IV (DPPIV), aminopeptidase A (APA) and gamma-glutamyl transferase (gamma-GT)], the S3 segment-specific marker intestinal type alkaline phosphatase (LAP) and as S1S2 marker (TN20-antigen), originally proposed as a surface marker for interstitial fibroblasts. Segment (proximal tubular vs. distal tubular) and proximal subsegmental (S1S2 vs. S3) expression of all five surface peptidases and TN20 antigen were first assessed by comparing immunohistochemical staining on normal human kidney tissue with staining for well-known segment-specific differentiation markers (intestinal type alkaline phosphatase, Tamm-Horsfall protein) on adjacent sections. All five peptidases were found to be expressed toa certain degree in all subsegments (S1 S2 and S3) of the proximal nephron, whereas expression was never seen int he more distal parts of the nephron. Flow cytometry was performed on cells obtained following gradient purification of collagenase-digested human renal tissue. Labeling cells for expression of LAP, NEP or DPPIV resulted in high yields of specifically labeled PTC (S1S2S3 origin). Labeling with anti-LAP resulted in the clearest distinction between positive and negative cell subpoulations, and therefore LAP was considered the best PTC marker for the use in FACS. LAP histochemical staining on sorted cells showed that flow sorting with monoclonal antibody (moAb) 250 (anti-intestinal type alkaline phosphatase) allowed sorting of S3 cells with > 90% purity. Likewise, moAb TN20 enabled us to obtain a highly purified S1S2 population as confirmed by the absence of LAP on sorted cells.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 25/11/20 alle ore 15:09:36