Figure 6 : Analysis of myosin heavy and light chains with DIA.

From: Mass spectrometry quantitation of proteins from small pools of developing auditory and vestibular cells

Figure 6

ah, Conventional myosin heavy chain genes over development using manual and automated data extraction. Fragment-ion and MS1 intensities were extracted from the manual and automated myosins dataset, averages for each peptide and sample condition were calculated across all samples. Only peptides detected in ≥8 samples were used for calculations. The intensity/average ratio was calculated for each peptide and each sample, and these were averaged across the three replicates for each sample condition. The average intensity/average across the three time points was calculated for GFP-positive and GFP-negative samples. Note that the trends were broadly similar for manual and automated extraction, as well as fragment-ion vs. MS1 intensities. g-l, Detection sensitivity for unconventional myosins in cochlear hair cells. Dilutions of a P0 GFP-positive cochlea sample were prepared (312, 625, 1250, and 2500 cells), and were analysed along with the standard 5000 cell samples. MS1 intensities were measured from DDA experiments (g,h) and DIA experiments (i–l) for selected myosins, and the sum of intensities was plotted for 2 (MYO3A and MYO3B) or 6 (MYO6 and MYO7A) peptides. Only unique peptides were used in the analysis. The data were fit with y=a1*x/(a2+x); for MYO3A, the final data point (asterisk) was not used in the fit. Each panel also includes the riBAQ (estimated mole fraction) for each protein from the DDA data. MYO3B was not detected in the DDA P0 GFP-positive cochlea samples.