Intact mass analysis by HRMS

Mass spectrometry (MS) is an analytical technique that measures the mass-to-charge ratio of charged particles.

  • Molecular weight determination
  •  Structural characterization
  • Qualitative and quantitative analysis of components in a mixture.

Areas of applications:

  • Proteomics
  • Pharmaceutical analysis
  • Forensic analysis
  • Environmental analysis
  • Clinical applications
Intact mass analysis by HRMS

ORBITRAP MASS SPECTROMETRY

Orbitrap mass spectrometers deliver a total possible maximum resolution (FWHM) of 1,000,000 at m/z 200 and a sub-1 ppm mass accuracy in a single compact and easy-to-use instrument. These high-resolution accurate-mass systems detect a wide range of compounds and small molecules during both targeted and untargeted analyses, without losing selectivity or sensitivity. Simply put, when it comes to Orbitrap technology, there is no compromise.

High Resolution Mass Spectrometer (HRMS) AT CBA FACILITY

Make: ThermoFisher Scientific

Model: Q Exactive Plus Biopharma Orbitrap Mass Spectrometer

Intact mass analysis by HRMS (2)
  • Resolution up to 240,000 (FWHM) at m/z 200
  • Analysis of large molecules up to m/z 8000

INTACT MASS ANALYSIS OF PROTEINS

  • Mass analysis of intact proteins using high resolution mass spectrometry is a rapid method for confirming the identity and the primary structure of proteins. 
  • Mass analysis can provide information for evaluating and comparing the isoforms, glycoforms, and molecule wide modifications associated with a protein. 
  • The analysis can readily compare manufacturing batches or can be used to compare a biosimilar to the innovator. 
  • Measurement of molecule weight is a requirement of the regulatory guidelines.

INTACT MASS –UNDER NON REDUCING CONDITIONS

Figure 2. Raw m/z spectrum of NIST mAb

Figure 3. Deconvoluted Mass (Da) spectra of NIST mAb

The charge envelopes in this spectrum are centered on ions having a charge (z) of 53. Each charge envelope is rich in information about the structure. The NIST mAb has four major glycoforms that can be observed in the inset of Figure 3.

INTACT MASS –UNDER REDUCING CONDITIONS

INTACT MASS –UNDER REDUCING CONDITIONS

INTACT MASS ANALYSIS AT CBA

 

 

 

Intact mass analysis of protein

Sample requirement

Amount- 0.1-0.5 ml (1mg/ml)

 

Deliverables

Raw mass spectrum and Deconvoluted mass spectra

Information required from Client

Expected molecular weight and formula

Sample solubility, polarity and formulation buffer 

Concentration

Protein sequence in FASTA format for comparison