英語翻譯
Results
Detection of upregulated proteins
In order to identify membrane proteins of K562 cells with
increased expression under iron deprivation,we isolated
hydrophobic membrane proteins from the cells after 24-h
incubation under control conditions (defined transferrin
medium supplemented with 5 lg/ml of transferrin as the
source of iron) and under iron deprivation (defined ironfree
medium without any added source of iron,see Materials
and methods) using a two-phase partitioning system
(2-D Sample Prep for Membrane Proteins from Pierce,see
Materials and methods).We prepared three pairs of 2D gels
from three separate pairs of samples of control cells and
cells cultured under iron deprivation.Gels were stained
using colloidal Coomassie blue.After scanning the stained
gels in a densitometer,we analyzed triplets of gel images
using PDQuest 7.1 2-D Analysis Software in semimanual
detecting and matching mode.
Approximately 300 stable spots were detected on each gel
map.The overall protein expression profiles with pI 3–11
and molecular mass of 20–120 kDa were very similar within
the three gels of both groups of samples (control and iron
deprivation).After the electronic construction of a virtual
Master gel,we searched for spots representing proteins with
significantly increased expression under iron deprivation
when compared with the expression under control conditions.
The increased expression under iron deprivation was
shown to be statistically significant in the case of two spots,
i.e.,spot 1 (P\0.05) and spot 2 (P\0.01).Positions of
these two spots on representative 2Dgels are shown in Fig.1
and the effect of iron deprivation on the expression of proteins
of these spots is shown in Fig.2.