title: Evaluation of Long-Term Cryostorage of Brain Tissue Sections for Quantitative Histochemistry.
abstract: Storage of tissue sections for long periods allows multiple samples, acquired over months or years, to be processed together, in the same reagents, for quantitative histochemical studies. Protocols for freezer storage of free-floating frozen sections using sucrose with different additives have been reported and assert that storage has no effect on histochemistry, but no quantitative support has been provided. The present study analyzed the efficacy of long-term storage of brain tissue sections at -80C in buffered 15% glycerol. To determine whether histochemical reactivity is affected, we analyzed 11 datasets from 80 monkey brains that had sections stored for up to 10 years. For processing, sections from multiple cases were removed from storage, thawed, and batch-processed at the same time for different histochemical measures, including IHC for neuronal nuclear antigen, parvalbumin, orexin-A, doublecortin, bromodeoxyuridine, the pro-form of brain-derived neurotrophic factor, and damaged myelin basic protein as well as a histochemical assay for hyaluronic acid. Results were quantified using stereology, optical densitometry, fluorescence intensity, or percent area stained. Multiple regression analyses controlling for age and sex demonstrated the general stability of these antigens for up to a decade when stored in 15% glycerol at -80C.
authors.affiliation: Department of Pharmacology and Experimental Therapeutics (LIE, AAR).
authors.forename: Larissa I
authors.initials: LI
authors.lastname: Estrada
authors.affiliation: Department of Pharmacology and Experimental Therapeutics (LIE, AAR).
authors.forename: Amy A
authors.initials: AA
authors.lastname: Robinson
authors.affiliation: Department of Anatomy and Neurobiology (ACA, ELG, NCH, FM, LBN, DER, RJK, DLR).
authors.forename: Ana C
authors.initials: AC
authors.lastname: Amaral
authors.affiliation: Department of Anatomy and Neurobiology (ACA, ELG, NCH, FM, LBN, DER, RJK, DLR).
authors.forename: Eustathia L
authors.initials: EL
authors.lastname: Giannaris
authors.affiliation: Department of Anatomy and Neurobiology (ACA, ELG, NCH, FM, LBN, DER, RJK, DLR).
authors.forename: Nadine C
authors.initials: NC
authors.lastname: Heyworth
authors.affiliation: Department of Anatomy and Neurobiology (ACA, ELG, NCH, FM, LBN, DER, RJK, DLR).
authors.forename: Farzad
authors.initials: F
authors.lastname: Mortazavi
authors.affiliation: Department of Anatomy and Neurobiology (ACA, ELG, NCH, FM, LBN, DER, RJK, DLR).
authors.forename: Laura B
authors.initials: LB
authors.lastname: Ngwenya
authors.affiliation: Department of Anatomy and Neurobiology (ACA, ELG, NCH, FM, LBN, DER, RJK, DLR).
authors.forename: Debra E
authors.initials: DE
authors.lastname: Roberts
authors.affiliation: Boston University School of Medicine, Boston, Massachusetts, and Department of Biostatistics, Boston University School of Public Health, Boston, Massachusetts (HJC).
authors.forename: Howard J
authors.initials: HJ
authors.lastname: Cabral
authors.affiliation: Department of Anatomy and Neurobiology (ACA, ELG, NCH, FM, LBN, DER, RJK, DLR).
authors.forename: Ronald J
authors.initials: RJ
authors.lastname: Killiany
authors.affiliation: Department of Anatomy and Neurobiology (ACA, ELG, NCH, FM, LBN, DER, RJK, DLR).
authors.forename: Douglas L
authors.initials: DL
authors.lastname: Rosene
mesh_terms: D000818:Animals; D034961:Antigens, Nuclear / Q000032:analysis; D001923:Brain Chemistry; D019208:Brain-Derived Neurotrophic Factor / Q000032:analysis; D001973:Bromodeoxyuridine / Q000032:analysis; D002452:Cell Count; D015925:Cryopreservation / Q000379:methods; D000089622:Doublecortin Domain Proteins; D005260:Female; D005629:Frozen Sections / Q000379:methods*; D006820:Hyaluronic Acid / Q000032:analysis; D007150:Immunohistochemistry / Q000379:methods; D008253:Macaca mulatta; D008297:Male; D008869:Microtubule-Associated Proteins / Q000032:analysis; D004676:Myelin Basic Protein / Q000032:analysis; D009419:Nerve Tissue Proteins / Q000032:analysis; D009479:Neuropeptides / Q000032:analysis; D000068797:Orexins / Q000032:analysis; D010320:Parvalbumins / Q000032:analysis
chemical_list: D034961:Antigens, Nuclear; D019208:Brain-Derived Neurotrophic Factor; D000089622:Doublecortin Domain Proteins; D008869:Microtubule-Associated Proteins; D004676:Myelin Basic Protein; D009419:Nerve Tissue Proteins; D009479:Neuropeptides; D000068797:Orexins; D010320:Parvalbumins; D006820:Hyaluronic Acid; D001973:Bromodeoxyuridine
keywords: batch-processing; brain tissue; cryopreservation; cryoprotection; frozen sections; immunohistochemistry; quantification; stereology