IBCN   20355
INSTITUTO DE BIOLOGIA CELULAR Y NEUROCIENCIA "PROFESOR EDUARDO DE ROBERTIS"
Unidad Ejecutora - UE
artículos
Título:
AN IMPROVED METHOD TO OBTAIN A SOLUBLE NUCLEAR FRACTION FROM EMBRYONIC BRAIN TISSUE
Autor/es:
GIUSTI, SEBASTIÁN; BOGETTI, MARÍA EUGENIA; BONAFINA, ANTONELA; FISZER DE PLAZAS, SARA
Revista:
NEUROCHEMICAL RESEARCH
Editorial:
SPRINGER/PLENUM PUBLISHERS
Referencias:
Lugar: Estados Unidos; Año: 2009 vol. 34 p. 2022 - 2029
ISSN:
0364-3190
Resumen:
This paper describes modifications of the standard methods for obtaining a soluble nuclear fraction
from embryonic brain tissue. The main improvements are: (1) the inclusion of a low speed centrifugation step
to prevent the appearance of high density contaminants, (2) a sucrose density gradient to remove
perinuclear mitochondria and ER membranes and (3) a protein extraction approach which significantly
enhances protein yield. To demonstrate the effectiveness of the method, pellets were analyzed by light and
electron microscopy and purity of the soluble extracts was immunologically tested. Finally, to illustrate
the applicability of this approach, the induction of the transcription factor HIF-1 (hypoxia-inducible
factor-1) was assessed by Western blot using soluble nuclear fractions and by immuno-electron microscopy
using purified nuclear fractions, both obtained from the optic lobes of chick embryos. In conclusion, the
procedure presently described appears to be reliable and convenient for obtaining a pure soluble nuclear
fraction from a discrete amount of embryonic brain tissue.