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. 2016 Mar 31;10:70. doi: 10.3389/fncel.2016.00070

Figure 2.

Figure 2

One dimension SDS-PAGE gel of secreted proteins precipitated from fibroblast conditioned-medium and conjugated with CyDyes. (A) Conjugation of CyDyes 3 and 5 to TCA-DOC and TCA-NLS-THF precipitated proteins. A 14% acrylamide/bisacrylamide (37.5:1) gel was used to resolve the CyDye conjugated supernatant protein. An internal standard was produced by conjugating protein from the TCA-DOC (Post-48 h) with both Cy3 (green) and Cy5 (red) dyes. One fibroblast cell line per tested condition for each of the studied pathology and control was tested and loaded in the gels in this order: pre-confluence/24 h, post-confluence/48 h, pre-confluence/48 h, post-confluence/48 h. Equal amount of proteins was loaded in all the gels. (B) Quantification of the fluorescent signal, detected with our laser-scanner Typhoon device, was done using ImageJ software. No statistical differences were noted for each of the tested condition, indicating that any of the protein precipitation method used in this study did not interfere with the CyDye conjugation.