Dendritic cell elimination as an assay of cytotoxic T lymphocyte activity in vivo.
We show in this paper that the survival of antigen-loaded dendritic cells in vivo may be used as a sensitive readout of CTL activity. We have previously shown that dendritic cells labeled with the fluorescent dye CFSE and injected sub-cutaneously into mice migrate spontaneously to the draining lymph node where they persist for several days. In the presence of effector CTL responses, dendritic cells loaded with specific antigen rapidly disappear from the draining lymph node. In this paper we extend the above observations and set up a simple and sensitive method to reveal CTL activity in individual mice in vivo. Dendritic cells were labeled with two different fluorochromes, loaded with antigen or left untreated, and mixed together before injection into mice. We show that only the dendritic cells loaded with specific antigen were cleared from the draining lymph node, while dendritic cells not loaded with antigen remained unaffected. Cytotoxic responses generated by immunization with peptide-loaded dendritic cells, or by infection with influenza virus, could be revealed using this method. Comparison of the differential survival of dendritic cells populations mixed together also allowed us to accurately evaluate the disappearance of dendritic cells, irrespective of variability in the injection site and other parameters. Given the ability of dendritic cells to efficiently take up and present complex antigens, nucleic acids and apoptotic bodies, this method may also allow the evaluation of cytotoxic activity against antigens that are not characterized in terms of peptide epitopes.
Ritchie
D S
DS
Malaghan Institute of Medical Research, Wellington School of Medicine, Wellington, New Zealand.
Hermans
I F
IF
Lumsden
J M
JM
Scanga
C B
CB
Roberts
J M
JM
Yang
J
J
Kemp
R A
RA
Ronchese
F
F
D000818:Animals; D000956:Antigens, Viral; D003602:Cytotoxicity, Immunologic; D003713:Dendritic Cells / Q000166:cytology / Q000276:immunology*; D018984:Epitopes, T-Lymphocyte / Q000276:immunology; D005260:Female; D005452:Fluoresceins; D005456:Fluorescent Dyes; D006023:Glycoproteins / Q000276:immunology; D015395:Histocompatibility Antigens Class I / Q000276:immunology; D008198:Lymph Nodes / Q000166:cytology / Q000276:immunology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D010446:Peptide Fragments / Q000276:immunology; D012235:Rhodamines; D013388:Succinimides; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014764:Viral Proteins
C116572:(((4-chloromethyl)benzoyl)amino)-tetramethylrhodamine; C087165:5-(6)-carboxyfluorescein diacetate succinimidyl ester; D000956:Antigens, Viral; D018984:Epitopes, T-Lymphocyte; D005452:Fluoresceins; D005456:Fluorescent Dyes; D006023:Glycoproteins; D015395:Histocompatibility Antigens Class I; D010446:Peptide Fragments; D012235:Rhodamines; D013388:Succinimides; D014764:Viral Proteins; C094244:glycoprotein peptide 33-41, Lymphocytic choriomeningitis virus