Introduction

Match-UP posts compares mass spec systems in the BioMS facility to give you an idea of how they compare. They are fact based but there will also be chunks of opinion and experience as well so these comparisons should be considered the beginnings of discussions with facility staff.

These comparisons assume you know about different MS technologies so please read this post first if you are not sure: https://sites.manchester.ac.uk/bioms-community/2020/04/08/basics-understanding-lc-ms-technologies/

Orbi Elite vs QE HF

Orbi Elite = nano-LC – nanospray – dual linear ion trap (LIT) – high field orbitrap

QE HF = nano-LC – nanospray – quadrupole – HCD cell – high field orbitrap

The two systems have a lot of similarity; they have the same LC, source and orbitrap portions. Where they differ is in the ion selection, fragmentation and the software used to control the systems. The QE HF is the newer system with newer technologies and software options. Lets consider the differences and how they will impact the analysis:

Ion selection: Quad vs LIT

This is a big difference. Linear ion trap (LIT) isolation means you need to fill the trap first before isolating the ions. In complex mixtures we are often interested in the minor components in the sample so these can be buried below much more abundant species. These abundant species will control the fill times in the trap and so ions that would need 100mS fill time may get <1mS and so never get above the level needed for detection and there is no way around this. Quadrupoles however continuously eject unwanted ions and therefore easily enrich for the low abundance molecules if you know what you want to look for. A quad system is much better at enriching low abundance species from high backgrounds (a quad infront of trap is even better – this is what you get in Thermo’s Tribrid systems).

Result: QE HF Wins

Fragmentation: HCD vs LIT

This is a more subtle difference. LIT fragmentation retains less stable ions such as b ions and so you get a much more information rich spectrum. However, the resolution and the mass accuracy of the spectrum is low – it is possible to scan the fragments in the orbitrap to get higher resolution and accuracy spectra but the Elite isn’t that optimised for this and it slows the system down a lot so this mode generally isn’t used. The HCD cell is more advanced and reliably fragments most molecules. It has some ion storage so the sensitivity is high. the fragments produced are predominately y ions so are easier to interpret. The Elite also has a HCD cell but it is an earlier version and in our hands it doesn’t work as well and the orbi is too slow to use it properly.

Result: Elite wins if you need the extra b ions for structural work but QE wins for everything else.

Technologies and software: New vs old

Although the orbitrap itself is the same in the two systems the technology around it is much newer in the QE HF and so it works faster and is possibly more sensitive. Changes in sensitivity can be hard to determine as both are ion traps and so offset sensitivity with increased fill times, you would see faster cycle times with more sensitivity due to shorter fill times but you may not see much difference in the MS signal. The biggest effect of this is in the MSMS as the changes mean that the QE HF can use the orbi for MSMS, giving higher mass accuracy and resolution data so you get many more matches.

Results: Not necessarily much of a win for the QE HF in MS1 but a big win in MS2.

In summary – QE HF wins…. in most cases but the Elite can still be great with a bit more effort

In complex mixtures the QE HF will be more sensitive and identify more peptides. This will be particularly clear in single samples where all identifications come from that run. In multiple sample sets the difference shrinks as identifications can be matched between runs. If you were only looking at MS1 data and using libraries of identified peptides then the difference would be slighter. The different form of fragmentation obtained can be beneficial in some cases and should be considered complementary is difficult samples. The QE HF remains our preferred system for virtually all sample types but you can still get good data out of the Elite if the QE queue is too big!

Tags: #Facility, matchups

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