Electrospray ionization (ESI) of intact proteins, peptides, and oligonucleotides produces mass spectra that contain series of multiply charged ions with associated mass-to-charge ratio (m/z) values. The resulting spectrum is complex and difficult to interpret, requiring mathematical algorithms for the analysis of the data.

Through a process called deconvolution, the BioPharma Finder application uses such algorithms to transform a charge state series into a molecular mass for an intact protein or oligonucleotide. To do this, the application identifies multiple peaks in the mass spectrum associated with different charge states of the same component and returns information about the masses and abundance of that component.

Intact Mass Analysis includes two independent deconvolution algorithms for mass spectral data:

  • Xtract, which deconvolves isotopically resolved mass spectra. See Xtract algorithm.
  • ReSpect™, which deconvolves isotopically unresolved (or unseparated) mass spectra. See ReSpect algorithm.

NOTE

Attempting to apply an algorithm to the wrong type of spectral data can lead to unreliable results.

In most cases, the Xtract algorithm fails to identify any components if you apply it to isotopically unresolved spectra as the components do not have isotopic profiles.

If you apply the ReSpect algorithm to isotopically resolved spectra, it might attempt to identify each isotopic peak as a separate component, rather than an isotopic composition of a single component.

For each of the two deconvolution algorithms, you can specify the deconvolution method:

  • Average Over Selected Retention Time: The application deconvolves and averages the user-specified source spectrum over a given retention time (RT) range.
  • Auto Peak Detection: The application generates the source spectra using the Parameterless Peak Detection (PPD) algorithm for the Auto Peak Detection of large molecules.
  • Sliding Windows: The application averages spectra over a succession of sliding windows in the RT range specified by the RT Range parameter. It deconvolves each of these averaged spectra and then merges similar masses to identify components.