title: In vivo labelling of the spleen with a red-fluorescent cell dye.
abstract: A method for labelling mouse spleen cells in situ is described. Spleen vessels were clamped before intrasplenic injection of a red-fluorescent cell dye (PKH-26). The labelling rate was 11.8% of all cells and 13.9% of B lymphocytes 30 min after injection as determined by FACS. 3 days later, the proportions of labelled cells were reduced to 7.4% (P < 0.01) and to 10.7% (P < 0.05), respectively. Only 10% of cells detected by FACS could be detected by fluorescence microscopy. Labelled cells were not found in peripheral lymph nodes 30 min after spleen injection, but were present 3 days later (FACS: 2.8% of all cells and 5.4% of B lymphocytes, P < 0.05 each), indicating migration of stained cells. Neither adverse nor toxic effects of in situ staining were observed. Isolated stained B lymphocytes from spleens of naive mice and from lymph nodes after immunisation with insulin showed proper function when tested in an ELISA-spot assay. The labelling procedure was used to follow splenic B lymphocytes producing natural anti-insulin antibody. These cells were found to be recruited for the early immune response in lymph nodes immunised with insulin.
authors.affiliation: Abteilung Klinische Endokrinologie, Medizinische Universitätsklinik, Freiburg, Germany.
authors.forename: A N
authors.initials: AN
authors.lastname: Khalaf
authors.forename: G
authors.initials: G
authors.lastname: Wolff-Vorbeck
authors.forename: K
authors.initials: K
authors.lastname: Bross
authors.forename: L
authors.initials: L
authors.lastname: Kerp
authors.forename: K G
authors.initials: KG
authors.lastname: Petersen
mesh_terms: D000818:Animals; D001402:B-Lymphocytes; D002465:Cell Movement; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D008198:Lymph Nodes / Q000166:cytology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008856:Microscopy, Fluorescence; D009930:Organic Chemicals; D013154:Spleen / Q000166:cytology*
chemical_list: D005456:Fluorescent Dyes; D009930:Organic Chemicals; C070080:PKH 26