Highly specific separation of heterogeneous cell populations by lectin-coated beads: application for the isolation of inner medullary collecting duct cells.
Conditions for the highly specific selection of a cell type by the use of lectin-coated magnetic beads are reported for the isolation of inner medullary collecting duct (IMCD) cells from a heterogeneous inner medullary cell suspension, containing both single cells and tubular fragments of variable size. The lectin Dolichos Biflorus Agglutinin (DBA), which binds in rat inner medulla exclusively to IMCD cells, was coupled via the avidin-biotin system to beads. By isolating DBA-bead-IMCD cells in a magnetic field (positive selection) from a suspension containing about 50% IMCD, a fraction of 98 +/- 1% purity was obtained; recovery of cells was up to 90%. Suspensions negative on reverse-transcriptase polymerase chain reaction for vimentin as a marker of contaminating interstitial and vascular cells could be received by repeating this procedure and additional trypsinization. On the other hand, it was possible to reduce the portion of IMCD cells in the suspension by one isolation step to 1.5 +/- 0.9% (negative selection). Performing this step twice resulted in virtually pure suspensions. No significant effects of this isolation technique on cell viability, growth characteristics, and biochemical parameters were observed. Therefore, this method appears to be a powerful tool for the highly specific separation of heterogeneous cell populations.
Grupp
C
C
Abteilung Nephrologie und Rheumatologie, Zentrum Innere Medizin, Georg-August-Universität, Göttingen, Deutschland. cgrupp@gwdg.de
Troche
I
I
Steffgen
J
J
Langhans
S
S
Cohen
D I
DI
Brandl
L
L
Müller
G A
GA
D000818:Animals; D002469:Cell Separation / Q000379:methods*; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D007679:Kidney Medulla; D007685:Kidney Tubules, Collecting / Q000166:cytology* / Q000378:metabolism; D037102:Lectins; D008863:Microspheres; D037121:Plant Lectins; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013997:Time Factors; D014746:Vimentin / Q000378:metabolism
D037102:Lectins; D037121:Plant Lectins; D014746:Vimentin; C036554:dolichos biflorus agglutinin