Confocal fluorescent intravital microscopy of the murine spleen.
Intravital microscopy has provided many insights into cellular interactions in various secondary lymphoid tissues. Because this technique allows for the visualization of cellular movement in real-time, it has been very powerful. However, until now, it has been difficult to apply this technique to the spleen. We report a technique that utilizes the Nikon RCM-8000 scanning laser, confocal microscope that allows for visualization of cellular movement in real-time in the rodent spleen. Using fluorescently labeled high molecular weight dextran or monoclonal antibodies, we are able to visualize fluorescently labeled cells rolling, tethering, and adhering in the spleen. In addition, we show that the majority of blood flow to the spleen remains within the white pulp nodules, as do most transferred erythrocytes at early time points. This is the first report of intravital microscopy of the spleen using a method that allows for easy identification of transferred cells.
Grayson
M H
MH
Division of Allergy and Immunology, Washington University School of Medicine, Campus Box 8122, 660 South Euclid Avenue, St. Louis, MO 63110, USA. wheeze@allergist.com
Chaplin
D D
DD
Karl
I E
IE
Hotchkiss
R S
RS
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002465:Cell Movement; D003911:Dextrans / Q000737:chemistry; D004912:Erythrocytes / Q000166:cytology; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives / Q000737:chemistry; D008214:Lymphocytes / Q000166:cytology; D051379:Mice; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000379:methods*; D013154:Spleen / Q000098:blood supply / Q000166:cytology*
D000911:Antibodies, Monoclonal; D003911:Dextrans; C015219:fluorescein isothiocyanate dextran; D016650:Fluorescein-5-isothiocyanate