Mass Spectrometry Identity Confirmation — ESI-MS vs. MALDI-TOF

Two mass spectrometry techniques dominate peptide identity confirmation: ESI-MS (electrospray ionization) and MALDI-TOF (matrix-assisted laser desorption/ionization, time-of-flight). This note compares them for QC use.

Ionization Principle

ESI-MS: Sample dissolved in volatile solvent passes through a charged capillary; the spray evaporates, leaving multiply charged peptide ions. Output is a charge-state envelope (e.g. [M+H]+, [M+2H]2+) requiring deconvolution.

MALDI-TOF: Sample co-crystallized with UV-absorbing matrix; laser pulse desorbs and ionizes peptides predominantly as [M+H]+. Time-of-flight separates by m/z.

Method Comparison

Attribute	ESI-MS	MALDI-TOF
Mass accuracy	≤5 ppm (Q-TOF/Orbitrap)	10–50 ppm
Charge states	Multiple	Single ([M+H]+)
Sample prep	LC-coupled, fast	Matrix co-crystallization
Best use	Online LC-MS QC	Quick offline ID

Use in Nexphoria QC

Nexphoria uses ESI-MS coupled to LC for routine identity confirmation. Observed [M+H]+ must match calculated monoisotopic mass within ±1 Da. Lot COAs report the observed mass.

RUO. Method note for documentation purposes.