Improvement of combined FISH and immunofluorescence to trace the fate of somatic stem cells after transplantation.
Fluorescence in situ hybridization (FISH) combined with immunohistochemistry of tissue-specific markers provides a reliable method for characterizing the fate of somatic stem cells in transplantation experiments. Furthermore, the association between FISH and fluorescent gene reporter detection can unravel cell fusion phenomena, which could account for apparent transdifferentiation events. However, despite the widespread use of these techniques, they still require labor-extensive protocol adjustments to achieve correct and satisfactory simultaneous signal detection. In the present paper, we describe an improvement of simultaneous FISH and immunofluorescence detection. We applied this protocol to the identification of transplanted human and mouse hematopoietic stem cells in murine brain and muscle. This technique provides unique opportunities for following the path taken by transplanted cells and their differentiation into mature cell types.
Donadoni
Chiara
C
Dipartimento di Scienze Neurologiche, Università di Milano, Padiglione Ponti, Ospedale Policlinico, Via Francesco Sforza 35, 20122 Milan, Italy. giacomo.comi@unimi.it
Corti
Stefania
S
Locatelli
Federica
F
Papadimitriou
Dimitra
D
Guglieri
Michela
M
Strazzer
Sandra
S
Bossolasco
Patrizia
P
Salani
Sabrina
S
Comi
Giacomo P
GP
D000818:Animals; D000831:Animals, Newborn; D016026:Bone Marrow Transplantation; D001921:Brain / Q000166:cytology / Q000601:surgery / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique / Q000379:methods*; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D051379:Mice; D008810:Mice, Inbred C57BL; D018482:Muscle, Skeletal / Q000166:cytology