Protein concentration was determined by the Bradford method using Bio-Rad Protein Assay with SmartSpec™ 3000 (Bio-Rad Laboratories, HerCAles, Candand Biotek μQuant (Winooski, VM).
Protein concentrations were measured by Bradford method using Bio-Rad Protein Assay (500 0006, Bio-Rad).
Protein concentration in the supernatants was determined by the Bradford dye method using Bio-Rad reagent.
Quantification of differential mRNA expression was calculated by the comparative Ct method using Bio-Rad CFX Manger 3.1 software.
The protein concentrations of the supernatants were determined by the Bradford method using BIO-RAD protein assay (BioRad) and normalized with lysis buffer.
The protein content in the lysates was measured according to Lowry method using Bio-Rad protein assay kit with bovine serum albumin as the standard.
Finally, RT-qPCR data were analyzed by the 2−ΔΔCT method using Bio-Rad CFX Manager Software.
After protein quantification by the Bradford method using the Bio-Rad Protein Assay reagent (Bio-Rad), proteins were boiled at 100°C for 5 min with 6x SDS (sodium dodecyl sulphate) sample buffer [350 mM Tris/HCl, pH 6.8, 10% (w/v) SDS, 30% (v/v) glycerol, 0.6 M DTT, 0.12 mg/ml bromophenol blue].
Protein concentration of the tissue extracts was determined by the Bradford method using the Bio-Rad Protein Assay system (Bio-Rad, Hercules, CA).
Then samples were sonicated and protein quantification was carried out according to the Bradford method, using a Bio-rad protein assay solution (Bio-rad, MI, Italy) and BSA as standard.
Phospho-EGFR (Tyr) was determined by a sandwich immunoassay method using a Bio-Plex phospho-EGFR (Tyr) assay kit (Bio-Plex phosphoprotein assay, Bio-Rad Laboratorins, Inc). in accordance with the manufacturer's instructions.
