A protocol for concurrent high-quality immunohistochemical and biochemical analyses in adult mouse central nervous system.
Eur J Neurosci
; 39(2): 165-75, 2014 Jan.
Article
em En
| MEDLINE
| ID: mdl-24325300
ABSTRACT
Biochemical analysis of central nervous system proteins and nucleic acids requires fresh-tissue homogenates, whereas immunohistochemistry usually is performed in sections prepared from perfusion-fixed tissue. Post-mortem immersion-fixation is possible, but largely impairs morphological preservation and protein antigenicity. Here, we present a simple, fast and versatile protocol allowing concurrent biochemical and immunohistochemical analysis, including pre-embedding immunoelectron microscopy, using tissue from the same animal. The protocol includes a brief transcardiac perfusion with ice-cold, oxygenated and glucose-supplemented artificial cerebrospinal fluid to maintain brain tissue alive, prior to isolation of regions of interest, followed by homogenisation for biochemistry or immersion-fixation for immunohistochemistry. We provide several examples demonstrating that this protocol allows optimal biochemical and morphological analysis, characterised with optimal sensitivity and preservation of tissue structure, along with a reduction of artefacts typically seen in perfusion-fixed tissue. This protocol should find widespread applications for combining analytical methods in tissue from the same animal, thereby reducing the number of mice required for a given experiment.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Encéfalo
/
Química Encefálica
/
Imuno-Histoquímica
Limite:
Animals
Idioma:
En
Revista:
Eur J Neurosci
Assunto da revista:
NEUROLOGIA
Ano de publicação:
2014
Tipo de documento:
Article
País de afiliação:
Suíça