Expert Insight: From Daltons to Megadaltons on a Single Mass Spectrometry Platform: Exclusive Webinar Highlights
26 Nov 2014
Discover the latest advancements in ultrahigh-resolution time-of-flight mass spectrometry and their applications to the analytical challenges facing innovators and biopharmaceutical/biosimilar characterization labs.
Dr Jason Wood, Market Area Manager for Bio/Pharmaceuticals, Bruker Daltonics, presented this helpful
webinar. A summary of the question and answer session is provided below. To watch the webinar on-demand,
click here.
1. My laboratory is in a GMP environment. Is the Bruker software 21CFR compliant?
Absolutely, Bruker’s compass software with security packing salt is compliant.
2. Why do you favor using the Fabricator enzyme over the traditional bottom-up approach? Fabricator has several advantages. Firstly, you can’t over-digest the antibody with the enzyme once it has cleaved within the hinge region, and secondly the enzyme doesn’t produce any strange artifacts. For example, in the case of trypsin, it might cleave the peptide multiple times so that you get small peptides that could get lost during HPLC separation. The Fabricator enzyme is excellent because you get defined pieces about 25 kDa in length and they are easy to analyze. So overall there are no artifacts, it is easy-to-use and you can’t over-digest the antibody.
3. Is the Fabricator enzyme readily available and where can I get it?
Fabricator is widely available and can be ordered from a distributor in your area.
4. How routinely can the maXis II determine the mono isotopic peak of the light and heavy chains, and what are the benefits of being able to do so? The
maXis II is routinely used to analyze and determine the mono isotopic peak of the light chain. You can actually obtain baseline resolution of the isotopes for the light chain, which is amazing. In terms of robustness, we have several instruments that have been running at customer labs and within Bruker, doing the sub-unit analysis. We are getting really good results and excellent baseline resolution for the light chain.
5. Much of the data you discussed focuses on top-down/middle-down approach, but my laboratory uses a bottom-up approach. What is the performance of the system for bottom-up? I did focus primarily on top-down and intact work, mainly because of the high resolution of the
maXis II allows us to analyze the larger proteins and heavy chains. However,
maXis II makes a great bottom-up machine. Pfizer does this for their peptide mapping in a pseudo bottom-up approach and they get great results.
6. Will the ETD approach be able to work like Thermo’s Orbitrap and do CID/ETD? Yes absolutely and it will also give better sequence coverage. We will present CID/ETD data in the future.
7. You didn’t say much about the new detector used in the maXis II; can you give any more details? It’s a new Bruker design and it is a multichannel plate detector with very fast single ion peak widths.
8. Is the analysis of deamidation in the middle-up approach routine, or is this a single lab’s experiment? This is a very routine way of doing this and we have several collaborators who are doing this at the moment; it is not a single lab’s experiment.
Watch the webinar on-demand Read this application note