Immunomagnetic selection of functional dendritic cells from human lymph nodes.
It was investigated whether positive immunomagnetic selection with two novel DC-specific mAb allowed purification of functional myeloid dendritic cells (MDC) and plasmacytoid dendritic cells (PDC) from the human lymph nodes (LN). The results were compared with enrichment of DC by low-density Nycodenz gradient centrifugation followed by immunomagnetic depletion of residual B- and T-cells (Nycodenz method). MDC were selected from inguinal LN cell suspensions using CD1c mAb and PDC using anti-BDCA-4 mAb. Immunomagnetic selection with anti-CD1c mAb yielded highly pure MDC preparations (90 +/- 3% MDC; n = 7), provided that the B-cells were thoroughly depleted by using CD19 magnetic beads and Large Depletion (LD) columns prior to selection of MDC. The purified MDC comprised both mature and immature cells and were functional, secreting large amounts of cytokines upon stimulation and strongly stimulating allogeneic T-cell proliferation. Immunomagnetic selection with anti-BDCA-4 mAb enriched PDC 70-fold to a purity of 59 +/- 26% (n = 8). The contamination consisted mainly of BDCA-4(+) T- and NK-cells. The previously used Nycodenz method yielded mixtures of MDC and PDC, not allowing functional studies of MDC and PDC separately. In conclusion, positive immunomagnetic selection with CD1c mAb from human LN cell suspensions yields almost pure MDC preparations, which are, in contrast to those obtained by the Nycodenz method, not contaminated with PDC. Moreover, these MDC are functionally intact. Selection with anti-BDCA-4 mAb does enrich PDC from human LN, but the resulting preparations are contaminated with T- and NK-cells.
Boor
Patrick P C
PP
Department of Gastroenterology and Hepatology, Room L-455, Erasmus Medical Center, Rotterdam, The Netherlands.
Ijzermans
Jan N M
JN
van der Molen
Renate G
RG
Binda
Rekha
R
Mancham
Shanta
S
Metselaar
Herold J
HJ
Kusters
Johannes G
JG
de Jong
Esther
E
Drexhage
Hemmo A
HA
Kwekkeboom
Jaap
J
D000911:Antibodies, Monoclonal / Q000276:immunology*; D018949:Antigens, CD1 / Q000276:immunology*; D000954:Antigens, Surface / Q000276:immunology; D002499:Centrifugation, Density Gradient; D016207:Cytokines / Q000096:biosynthesis; D003713:Dendritic Cells / Q000276:immunology* / Q000378:metabolism; D005434:Flow Cytometry; D006023:Glycoproteins / Q000276:immunology*; D006801:Humans; D018189:Immunomagnetic Separation; D016130:Immunophenotyping; D007472:Iohexol; D008198:Lymph Nodes / Q000166:cytology / Q000276:immunology*; D022423:Myeloid Cells / Q000276:immunology* / Q000378:metabolism; D010950:Plasma Cells / Q000276:immunology
D000911:Antibodies, Monoclonal; D018949:Antigens, CD1; D000954:Antigens, Surface; C437944:CD1C protein, human; D016207:Cytokines; D006023:Glycoproteins; C478196:blood dendritic cell antigen 4, human; D007472:Iohexol