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Post-translational modification (PTMs) which are most characteristic in protein biopharmaceuticals include glycosylation (which comprises galactosylation, fucosylation, high mannose derivatives and sialylation), oxidation, phosphorylation, sulfation, lipidation, disulfide bond formation and deamidation, as well as (to a more limited extent) carboxylation, hydroxylation, and amidation; with many biopharmaceuticals having a combination of two or more PTMs. PTMs need to be monitored during numerous stages of the manufacturing process and require identification, control of their levels, and assessment of their impact on the protein. In order to do this many chromatographic (reversed phase and HILIC HPLC) and mass spectrometric (MS) techniques are employed. 

This webcast will consider the types of modifications which can be assessed and what methods should be used to identify them, how various MS scanning techniques can be used for PTM analysis, as well as considering if the analysis mode itself can induce post translation modifications (collision induced dissociation vs. electron transfer dissociation (ETD) and electron capture dissociation (ECD)). Finally, the use of LC-MS for protein determination of antibody-drug conjugates in biological matrices will be discussed.

 

Topics include:

  • What post translational modifications can be assessed?
  • What effect do post translation modifications have on protein biopharmaceuticals?
  • Reversed phase HPLC analysis of PTMs
  • Utilizing different MS scanning functions to enable post translational modification events
  • How the analysis method can alter the post translation modification - alternatives to classical collision induced dissociation (top down analysis)
  • Host cell protein analysis using LC-MS
  • Protein quantification using LC-MS (pharmacokinetic studies) - protein determination, in particular ADC determination, in biological matrices