Journal: Molecular & Cellular Proteomics

Article Title: Identification of Differentially Regulated Secretome Components During Skeletal Myogenesis

doi: 10.1074/mcp.m110.004804

Figure Lengend Snippet: FIG. 2. 1D-SDS PAGE: A, for examining the incorporation of [13C6]- lysine, B, for determining the thresholds for differential expression, and C, for secretome analysis during skeletal myogenesis. M shows the protein ladder.

Article Snippet: One-dimensional (1D)-SDS-PAGE—All 1D-SDS PAGE were performed on 12%-polyacrylamide gels (10 cm 10 cm, Bio-Rad) using Coomassie blue (ICN Biomedicals, Inc., Costa Mesa, CA) to visualize the resolved protein bands.

Techniques: SDS Page, Quantitative Proteomics

Journal: Molecular & Cellular Proteomics

Article Title: Identification of Differentially Regulated Secretome Components During Skeletal Myogenesis

doi: 10.1074/mcp.m110.004804

Figure Lengend Snippet: FIG. 5. Comparison of SILAC and Western blot analyses. In SILAC, samples from set-1 and -2 were analyzed with Agilent 1100 RPLC-LTQ LIT tandem mass spectrometer and LC Packings RPLC-QSTAR Pulsar hybrid QqTOF tandem mass spectrometer respectively. Peptide ratios were determined by the ratio of heavy- versus light-labeled monoisotopic peak intensity, as indicated by red- and green-dashed lines in the representative MS mass spectra. Average and S.D. of which contributed to the corresponding protein ratios (H:L) and error respectively as revealed beneath each MS mass spectrum. For verification, the same set of samples were subjected to Western blot analyses twice as shown in the column “Western blot-1 ” and “Western-blot-2 ” under the “serum-free SILAC culture.” Protein ratios were deduced by the ratio of fluorescence intensity associated with the protein band in MT- versus MB-lane (MT:MB) as revealed beneath each blot. Protein ratio of “9999” and “0” represented the protein was exclusively expressed in MT and MB respectively. To verify the protein expression levels in the classical culture conditions, i.e. 2% horse serum (HS), replicate sets of conditioned media derived from the 2% HS culture condition were subjected to Western blot analyses, and the results were tabulated under the column “2% HS culture.” Coomassie blue staining of the 1D-SDS PAGE was used as the loading control.

Article Snippet: One-dimensional (1D)-SDS-PAGE—All 1D-SDS PAGE were performed on 12%-polyacrylamide gels (10 cm 10 cm, Bio-Rad) using Coomassie blue (ICN Biomedicals, Inc., Costa Mesa, CA) to visualize the resolved protein bands.

Techniques: Comparison, Multiplex sample analysis, Western Blot, Mass Spectrometry, Labeling, Fluorescence, Expressing, Derivative Assay, Staining, SDS Page, Control