LC-MS/MS Analysis of the Proteomic Profile of Neuromelanin Granules Isolated from Human Brain Tissue

Begin with peptides derived from neuromelanin granules (NMGs) isolated from postmortem human brain tissue.

NMGs, found in dopamine-producing neurons, contain neuromelanin pigment, proteins, and lipids.

Take a high-performance liquid chromatography (HPLC) column containing a silica-based stationary phase functionalized with hydrophobic ligands.

Load the peptides onto the column.

Hydrophobic peptides bind more strongly to the column than hydrophilic ones.

Pass an elution buffer through the column.

Less hydrophobic peptides elute first, while more hydrophobic ones elute later.

The eluted peptides enter the mass spectrometer's electrospray ionization source, where they are ionized into charged peptide ions.

The ionized peptides then enter the mass analyzer, where they get separated based on their mass-to-charge ratios.

A selected peptide ion is directed to the collision cell and fragmented into smaller ions, whose mass-to-charge ratios are measured again.

Finally, the data is compared to a reference database to identify the proteins present in the NMG sample.