Bottom-up proteomics digests a protein with trypsin (cuts after K/R residues) into peptides, ionises them by electrospray, and separates the ions in a time-of-flight (TOF) tube: lighter m/z ions arrive at the detector first.
t = L · √(m / (2·z·e·V))
t: flight time L: tube length m: ion mass
z: charge state e: electron charge V: accelerating voltage
- Protein length / missed cleavages — regenerates the tryptic peptide list (more missed cleavages = fewer, longer peptides), shown as coloured ions entering the flight tube.
- Ion injection rate — how many peptide ions are launched into the analyser per second.
- Analyser resolution — how tightly ions of similar m/z are resolved into separate peaks on the detector trace (real instruments trade resolution for scan speed).
- Show DB matches — highlights ions whose mass matched a peptide predicted from the reference protein (green) vs unmatched background (grey), the core idea behind database search and false-discovery-rate (FDR) filtering.
Real-world relevance: this is exactly how instruments like Orbitrap and TOF mass spectrometers identify thousands of proteins in a single sample, underpinning modern proteomics and biomarker discovery.