After incubation of the cells in blocking solution with donkey anti-mouse IgG Cy3
conjugated secondary antibody (1:500) (Millipore, USA) for 2 h at room temperature in the dark, the cells were washed with PBS.
and Cy5) with excitation in the visible and near-infrared or infra-red region.
Briefly, 120 pg of total protein from each exposed sample and from each control sample was labeled with 400 pmol of the cyanine dyes Cy5 and Cy3
(GE Healthcare), respectively.
The concentration of the purified samples and Cy3
dye incorporation efficiency was evaluated using a NanoDrop 1000 spectrophotometer.
5 [micro]g/mL monoclonal mouse antivimentin conjugated to Cy3
(Sigma-Aldrich, clone V9).
On array 1 and array 3, the exercised muscles were incorporated with Cy3
dye, and on array 2 and array 4, exercised muscles were incorporated with Cy5 dye.
leprae strains (One reference strain and another obtained from clinical samples) by comparative genomic hybridization according to the protocol already standardized in our laboratory Cy 5 labelled DNA of standard strain Thai-53 and Cy3
labelled DNA of a lepromatous leprosy patient was used in the microarray experiments for identification of loci showing variability.
For microarray analysis, total RNA was isolated, labeled with Cy3
, and hybridized to Agilent Human Whole Genome microarrays.
In the laboratory, each sample is labeled using red Cy5 and green Cy3
The use of an array of serial dilutions of the Cy5 and Cy3
dyes with many of the characteristics of an ideal material was explored to measure scanner performance over a 5-week period.
After isolation of mRNA using Trizol, mRNA from two sources (for example 4th generation 2,4-D and 4th generation control) was labeled with Cy5 and Cy3
using PCR and hybridized to a Drosophila microarray.
This technique involved pre-labeling two protein extracts, using two fluorescent cyanine dyes known as Cy3