title: Isolation, propagation, and analysis of biological nanoparticles.
abstract: Calcifying biologic nanoparticles (NPs) have been implicated as nucleation points for a number of -pathologic events that include vascular calcification and the formation of kidney stones. In order to study these potential relationships, reproducible isolation of well-characterized biologic NPs is a necessity. Our group has isolated and propagated calcifying NPs from several human tissues and renal stones. Specific proteins that could nucleate a calcium phosphate shell under physiologic conditions have been identified as part of their structure, including elongation factor Tu (EF-Tu) and fetuin-A. Visualization, using advanced transmission electron microscopy (TEM), immunofluorescence microscopy, and nuclear and antibody staining in conjunction with flow cytometry, can further elucidate NPs composition and their role in pathology. In order to allow uniform investigation by others, the isolation, culture, and handling procedures for biologic NPs from human calcified vascular tissue and kidney stones are reported in detail.
authors.affiliation: Division of Nephrology and Hypertension, Department of Medicine, Mayo Clinic, Rochester, MN, USA.
authors.forename: Michael P
authors.initials: MP
authors.lastname: Linnes
authors.forename: Farooq A
authors.initials: FA
authors.lastname: Shiekh
authors.forename: Larry W
authors.initials: LW
authors.lastname: Hunter
authors.forename: Virginia M
authors.initials: VM
authors.lastname: Miller
authors.forename: John C
authors.initials: JC
authors.lastname: Lieske
mesh_terms: D015153:Blotting, Western; D002114:Calcinosis / Q000378:metabolism; D002129:Calcium Oxalate / Q000378:metabolism; D002130:Calcium Phosphates / Q000378:metabolism; D005591:Chemical Fractionation / Q000379:methods*; D002725:Chloroform / Q000737:chemistry; D004247:DNA / Q000737:chemistry / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D007669:Kidney Calculi / Q000378:metabolism; D013058:Mass Spectrometry; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D053758:Nanoparticles / Q000032:analysis* / Q000737:chemistry*; D019800:Phenol / Q000737:chemistry
chemical_list: D002130:Calcium Phosphates; D002129:Calcium Oxalate; D019800:Phenol; D002725:Chloroform; D004247:DNA; C020243:calcium phosphate