title: Scanning electron microscopy preparation protocol for differentiated stem cells.
abstract: The lack of an established protocol for scanning electron microscopy (SEM) studies on stem cells differentiating into adipogenic lineage led us to develop a protocol for the preparation of differentiated adult bone marrow-derived mesenchymal stem cells (BMSC) for SEM. This protocol describes the procedure to maintain and preserve the structural organization of cellular components following differentiation, for morphological and physical characterization. The fixation of the differentiated cells was followed by dehydration using methanol, and vacuum desiccation before microscopy. The use of longer chain alcohols as dehydrating agents was avoided in our method to reduce the dissolution of lipid deposits in cells, thus allowing the maintenance of their structural integrity. The time period for the processing of samples was reduced by avoiding the osmium tetroxide postfixation and critical point drying. Thus, this protocol helps in determining the potential, fate, and degree of stem cell differentiation. This may be useful for SEM analysis of differentiated cells, especially those grown on various scaffolds.
authors.affiliation: Stem Cell and Molecular Biology Laboratory, Department of Biotechnology, Indian Institute of Technology Madras, Chennai 600 036, TN, India.
authors.forename: Sreejit
authors.initials: S
authors.lastname: Parameswaran
authors.forename: Rama S
authors.initials: RS
authors.lastname: Verma
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology; D002454:Cell Differentiation; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D051379:Mice; D008855:Microscopy, Electron, Scanning / Q000379:methods*; D009993:Osmium Tetroxide / Q000737:chemistry
chemical_list: D009993:Osmium Tetroxide