Antibodies against macrophages that overlap in specificity with fibroblasts.
BACKGROUND
Fibroblasts can be misidentified as macrophages because both cell types share antigens that are associated with popular antibodies targeting the monocyte/macrophage lineage. With the recent description of fibroblast-specific protein 1 (FSP1), we revisited the specificity of antibodies directed against macrophages to determine systematically which antibodies best distinguish both cell types in fibrotic tissues.


METHODS
Tissue fibrosis was produced in mice carrying the GFP transgene encoding green fluorescent protein under the control of the FSP1 promoter. Single cell suspensions from these marked tissues were submitted for flow cytometry using antibodies against Mac-1, Mac-2, Mac-3, F4/80, CD68, major histocompatibility complex (MHC) class II, and CD45, and cDNA amplification of mRNA encoding the above target antigens was performed using specific primer sets in sorted pools of cells. Fibrotic tissues were also stained by immunohistochemistry with the same antibodies and examined under confocal microscopy.


RESULTS
Comparison overlap between FSP1(+) fibroblasts with each of the macrophage markers demonstrated that all antimacrophage antibodies (Mac-1, Mac-2, Mac-3, CD68, MHC class II, and CD45) except one (F4/80) recognize both cell types.


CONCLUSIONS
Antibodies directed against F4/80 clearly distinguish macrophages from FSP1(+) fibroblasts in fibrotic tissues and is the preferred antibody in mice.
Inoue
Tsutomu
T
Department of Medicine, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
Plieth
David
D
Venkov
Christo D
CD
Xu
Carol
C
Neilson
Eric G
EG
D000818:Animals; D000918:Antibody Specificity; D015703:Antigens, CD / Q000032:analysis; D000943:Antigens, Differentiation / Q000032:analysis; D015214:Antigens, Differentiation, Myelomonocytic / Q000032:analysis; D002135:Calcium-Binding Proteins / Q000032:analysis; D005347:Fibroblasts / Q000276:immunology*; D005355:Fibrosis; D037502:Galectin 3 / Q000032:analysis; D007668:Kidney / Q000473:pathology; D016177:Macrophage-1 Antigen / Q000032:analysis; D008264:Macrophages / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic; D000071999:S100 Calcium-Binding Protein A4; D009418:S100 Proteins; D000097382:CD68 Molecule
D015703:Antigens, CD; D000943:Antigens, Differentiation; D015214:Antigens, Differentiation, Myelomonocytic; D002135:Calcium-Binding Proteins; D037502:Galectin 3; D016177:Macrophage-1 Antigen; D000071999:S100 Calcium-Binding Protein A4; D009418:S100 Proteins; C081026:CD68 protein, mouse; D000097382:CD68 Molecule; C470944:S100a4 protein, mouse; C036763:monocyte-macrophage differentiation antigen