Post on 10-Jul-2020
Incorporation of FAIMS into LC-MS
‘omics’ analysis
Kayleigh L. Arthur, Matthew A. Turner, James C. Reynolds and Colin S. Creaser Centre for Analytical Science, Department of Chemistry, Loughborough University, Loughborough, LE11 3TU, U.K.
LC-MS
LC-FAIMS-MS
Chromatographic
Peak
Compensation
Field scan
– 11 CFs
Mass Spectra
6 s
1 s
Figure 4: Representation of LC-FAIMS-MS peak and
scan timescale and peak deconvolution for each CF
KAYLEIGH ARTHUR
Centre for Analytical Science, Department of Chemistry, Loughborough University
K.Arthur@lboro.ac.uk, uk.linkedin.com/kayleigh-arthur-0512348a
• In non-targeted ‘omics’ experiments such as metabolomics and proteomics,
typically liquid or gas chromatography (LC or GC) combined with mass
spectrometry (MS) is used to separate and analyse complex biological
matrices
• Molecular features can however be missed or remain hidden within the
dataset using these techniques due to:
o Trace level features unresolved from the noise
o Unresolved isomeric or isobaric species
• We propose the use of field asymmetric waveform ion mobility spectrometry
(FAIMS) in a scanning capacity (Figure 2) in conjunction with LC-MS to
improve peak capacity and reduce chemical noise
Agilent
6230
TOF MS
Drying gas flow
Spray shield
Miniaturised
FAIMS
Inlet capillary
Nebuliser Sheath
gas flow
Nozzle
Liquid flow
Figure 1: Schematic diagram
of the modified Agilent
JetStream electrospray
source with miniaturised
FAIMS device
• The data presented here used a miniaturised
FAIMS chip (ultraFAIMS, Owlstone Ltd.) located
in the modified ion source region of a time-of-
flight-MS (Agilent 6230 TOF), in front of the MS
inlet capillary (Figure 1)
• FAIMS is a gas phase ion
separation technique that
utilises an oscillating high
frequency (RF) waveform,
known as the dispersion
field (DF)
• Rapid separation of gas
phase ions is a result of
differences in an ion’s
mobility in a buffer gas
under alternating high and
low electric fields (Figure 2)
Figure 2: Representation of ion transmission through
planar FAIMS electrodes
• A small DC voltage known as the compensation field (CF) can be superimposed
on the DF in order to selectively transmit ions, or scan through CF values to
transmit ions • Separation based on ion mobility makes FAIMS
orthogonal to both LC and MS
• The main limitation to combining FAIMS with LC-
MS for scanning experiments is scanning the
FAIMS on a timescale compatible with a
chromatographic peak
Miniaturised chip-
based FAIMS in chip
housing 700 µm
100 µm
Side view of
FAIMS chip
Parallel pairs
of electrodes
To MS N2
Figure 3: Miniaturised FAIMS
device
• The fast scanning capabilities of
the miniaturised FAIMS chip
(Figure 3) allows an entire FAIMS
scan to be acquired per second,
allowing enough data points per
LC peak (Figure 4)
• Benefits from using LC-FAIMS-MS analysis compared to LC-MS:
o Trace level features can be separated from chromatographic noise
o Isomeric or isobaric species can be resolved at different CFs
o Improved peak capacity
• FAIMS conditions for use in LC-
FAIMS-MS analysis chosen from the
direct infusion of human urine into
FAIMS-MS (Figure 5)
• DF 240 Td, CF scanned between -0.9 –
4 Td at 0.49 Td step sizes
Figure 5: FAIMS-MS DF vs CF
plot of human urine infusion
Figure 6: LC-FAIMS-MS heat map of total
ion chromatograms at each CF
• From the deconvoluted total ion
chromatograms (Figure 6) can
already see multiple examples of
peaks at the same retention time
that appear in multiple FAIMS CFs,
but we can look closer at
individual m/z values (Figure 7)
• Ions at the same m/z and same retention
time can be separated using FAIMS
Figure 7: LC-FAIMS-MS heat map of
m/z 359.316
3 x10
0
1
2
3
Counts vs. Retention Time (min)
5.1 5.2 5.3
4 x10
0
1
2
3 5.212
3 x10
0
1
2
3 5.217
3 x10
0
0.5
1 5.203
Counts vs. Retention Time (min) 4.4 4.6 4.8 5 5.2 5.4 5.6 5.8 6
LC-MS
FAIMS off
LC-FAIMS-MS CF 0.08 Td
LC-FAIMS-MS CF 2.04 Td
Figure 8: LC-FAIMS-MS chromatogram
comparison with LC-MS, of m/z 207.11 at
retention time 5.21 min
m/z 207.111
RT 5.21 min
FAIMS scanned
from CF -0.9 to 4 Td
LC-FAIMS-MS
FAIMS off = 1 peak, FAIMS on = 2 peaks
• Figure 8 shows an example of the
separation of isobaric species
(same m/z and RT) using LC-
FAIMS-MS and an increase in peak
capacity
• Figure 9 shows an example of the
reduction of chemical noise using
LC-FAIMS-MS so that the peak S:N
is improved using FAIMS
3 x10
0
2
4
3 x10
0
1
2
Counts vs. Retention Time (min)
0.5 1 1.5 2 2.5 3 3.5 4 4.5 5 5.5 6 6.5 7 7.5 8 8.5 9 9.5
LC-MS FAIMS off
LC-FAIMS-MS
CF 0.08 Td
m/z 331.210, RT 4.24 min; FAIMS off = 0 peak, FAIMS on = 1 peaks
FAIMS off S:N = 2.7
FAIMS on S:N = 124.1
Figure 9: LC-FAIMS-MS chromatogram
comparison with LC-MS, of m/z 331.21 at
retention time 4.24 min
ACKNOWLEDGEMENTS
The authors thank Owlstone Ltd., Agilent Technologies and Loughborough University for financial and technical support
The authors thank the Royal Society of Chemistry for the award of a bursary for conference attendance