Intact Method

During ionization, a purified intact molecule forms charged molecular species rather than being described only through fragments. The mass spectrometer then measures each species’ mass-to-charge ratio, providing the measurements used to determine molecular mass. Because the whole protein remains represented, this readout can support direct identity confirmation and comparison of molecular forms.

Mass-to-charge measurements allow the instrument to determine the molecular mass of the analyzed biomolecule. Differences in the resulting mass can reveal changes such as post-translational modifications, which are chemical alterations to a protein after its production. This makes the measurement useful for examining molecular composition without relying exclusively on information from digested fragments.

The Intact Method preserves the complete molecule, whereas peptide-based workflows examine fragments produced after digestion. Fragmentation can provide detailed information about parts of a protein, but it may obscure the composition of the original whole molecule. Using both strategies can therefore combine direct molecular-level assessment with information obtained from peptide analysis.

The workflow begins with a purified protein or other biomolecule sample. The sample is introduced into a mass spectrometer, where ionization produces charged molecular species. The instrument measures their mass-to-charge ratios, and those measurements are used to determine molecular mass. The resulting analysis can then support identity confirmation, heterogeneity assessment, or quality control.

Intact analysis can detect changes such as post-translational modifications by comparing the molecular mass measured for the complete protein. It can also expose sample heterogeneity, meaning the presence of multiple molecular forms within a preparation. These outcomes help investigators evaluate whether a sample contains the expected composition and whether distinct forms may require further examination.

Researchers can apply the approach to protein identity confirmation, sample heterogeneity assessment, and quality control. In biochemistry, it provides a direct view of molecular composition for purified biomolecules. In biopharmaceutical research, retaining the whole protein supports evaluation of product-related molecular forms, complementing peptide-based analyses when confirmation of the complete molecule is important.