Page 21 - Shimadzu Journal vol.9 Issue1

Clinical Research

In contrast, PESI-MS omits chromatography by directly captur- was immediately centrifugated, while the second was delayed
ing tiny sample amounts with a needle tip from which electrospray for 3 h.
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is generated in ambient conditions by applying a high voltage . Plasma aliquots were thawed in a water ice bath (0°C) for 15
This simplifies handling immensely, shortens measurement times min. 10 µl of each plasma sample aliquot was pooled for quality
to minutes and allows previously impossible application scenar- control (QC) and split into 20 µl aliquots. From each sample, QC
4–6
ios , especially where robust, routine and high-throughput meas- aliquot and blank a volume of 20 µl were precipitated with 380
urements are needed. Sample quality control is one such setting µL extraction solution to a final concentration of 10 mM NH4Ac,
where metabolomics has shown great scientific promise but has not 70% MeOH and 5% DMSO. Precipitates were frozen at -80°C
yet entered routine application. until measurement within 3 h after thawing of samples. The meas-
High-quality biospecimens are crucial for reproducible results urement was split into 3 batches. Blanks and QC were measured
in *omics or biomedical research, for correct clinical routine di- repeatedly throughout the sequence. Precipitated extracts were
agnostics and for successful biobanking. Currently specific types thawed in 1–2 h sub-batches in water ice baths and precipitates
of preanalytical issues are tested with specific methods . These were removed by centrifugation for 5 min at 4°C with 12.000 rpm.
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specific methods are not comprehensive enough to cover the most Supernatants were kept in a water ice bath until measured
frequently occurring pre-analytical error sources in one swift meas- with the DPiMS-2020 (Shimadzu Corporation, Kyoto, Japan). Per
urement. NMR and MS-based metabolomics has shown great measurement 10 µl were deposited on a sample plate and all meas-
capability to detect various pre-analytical errors in one measure- urements were replicated until at least two valid TIC patterns for
ment 8–14 . However, costs and complexity precede a wide-spread each ionization mode were recorded. TIC patterns were defined
routine application, despite the numerous benefits of metabolomics as invalid when TIC spiking stopped before at least 30 s of the
for sample quality control. mode were measured (refer to Fig. 1 for invalid TIC pattern ex-
PESI-MS could overcome this barrier through its unique ad- ample). For instrument settings refer to Table 1. For each replicate
vantages and capability to deliver full mass spectra within minutes. a fresh needle (silicon coated, 18529A1, Shimadzu Corporation,
In our set-up PESI was coupled to a single-quad mass spectrome- Kyoto, Japan) and sample plate (11A9722418115OMS, Shimadzu
ter, purposely omitting and more sophisticated detectors in order Corporation, Kyoto, Japan) was used.
to (1) decrease instrument acquisition and maintenance costs, (2)
increase the instruments robustness and operating usability for per-
Table 1. Instrument settings for the PESI measurement.
sonnel without extensive MS expertise, and (3) decrease measure-
IONIZATION
ment time per sample.
In this proof-of-principle study we concentrated on the single, Needle position -37 mm
most common preanalytical issue, the time delay of blood to Outage time 200 ms
plasma processing. Our aim was to investigate if PESI-MS is SAMPLE TAKE
able to robustly classify time delays of 3 h from immediately pro-
Needle position -46 mm
cessed sample.
Outage time 50 ms
Speed 250 mm/s
MEASUREMENTS
Materials & Methods voltage segment Electric potential Time
1 – Corona +4.00 kV 3.6 s
Plasma preparation and PESI-MS measurement 2 – neg mode -4.25 kV 30.0 s
The study was conducted in adherence to the Declaration of 3 – Corona -4.50 kV 3.6 s
Helsinki and was reviewed by the ethical committee of the Medical
4 – pos mode +2.75 kV 30.0 s
University of Graz, Austria (31–116 ex 18/19, 16.01.2019). Study details
m/z range 10-2000 m/z
are available at https://www.drks.de DRKS-ID: DRKS00024807.
Scan speed 5000 u/s
Blood samples were donated by 50 volunteers (24 female, 26 male,
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age 18–90, BMI≥ 18.5 kg/m ) and were collected within 3 weeks (for
15
details refer to Bordag et al 2021 ). From each volunteer one sample

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