FIGURE 2: Video frames capture the acoustic droplet ejection process (TOP). Summary of ADE absolute accuracy (< 10%) and precision (< 8% CV) showing consistent quality across fluid classes (BOTTOM).

Reference: 1. Van Berkel, G.J., Kertesz, V. Rapid Communications in Mass Spectrometry 2015 29 (19) pp 1749-1756 DOI: 10.1002/rcm.7274

FIGURE 2: Photo of the initial ADE-OPP setup. We operate with the OPP oriented “upside down” to capture droplets ejected upward from the fluid meniscus of a multi-well coupon.

FIGURE 6: 10 nanoM reserpine in 50:50 Methanol:Water droplets ejected by ADD to OPP operating in vortex mode. Number of droplets varies from 1 – 200 (2.5 – 500 nL). Droplet ejection repetition rate was adjusted to keep total injection time constant at 1 second (TOP).

Linearity of peak area vs. number of droplets, R2 = 0.9975. (BOTTOM).

FIGURE 5: Signal is dependent on the size of the hanging droplet. Larger droplet volume leads to higher dilution and broader, lower intensity peaks.

FIGURE 7: Single droplet injection series, 100 nanoM reserpine in 50:50 MeOH:Water (TOP) and droplet ladder from 1 to 10 droplets (MIDDLE). Droplet ladder repeated with analyte matrix containing 90% digested plasma (BOTTOM).

FIGURE 8: Single droplet injection series, 100 nanoM reserpine in 50:50 MeOH:Water. For single droplet injection, no matrix effects observed up to pure digested plasma.

FIGURE 4: Details of the OPP sampling end (area in dotted circle) are shown enlarged. Liquid meniscus type: (1) supercritical vortex, (2) critical vortex, (3) subcritical vortex, (4) balanced, and (5) hanging drop. Meniscus type is determined by relative values of the solvent flow rate into the probe annulus and the solvent aspiration rate though the central capillary to the ionization source. Arrows indicate basic solvent flow paths.

FIGURE 3: Open port probe oriented to capture droplets ejected by ADE.

A New Platform for High-Throughput Mass Spectrometry:Acoustic Droplet Ejection with an Open Port Probe Sampling InterfaceLucien Ghislain 1, Chang Liu 2, Don Arnold 3, Sammy Datwani 1 1 Labcyte Inc., San Jose CA, USA; 2 Sciex, Concord, Ontario, Canada; 3 Veristad LLC, Livermore, CA

© 2018 LABCYTE INC. All rights reserved. Labcyte® and Echo® are registered trademarks of Labcyte Inc., (“Labcyte”) in the United States and/or other countries.POS-ACMS-1.0

SLAS 2018

IntroductionWe demonstrate a system combining acoustic droplet ejection (ADE) with the open port probe sampling interface for high-throughput mass spectrometry (MS). We use a customized Labcyte Echo® acoustic liquid handler to eject 2.5 nL droplets directly from a 384-well assay plate into an Open Port Probe (OPP) sampling interface. Captured droplets are diluted into a continuous flow of solvent and the OPP fluid stream is aspirated into the MS electrode where sample is ionized by standard electrospray prior to entering a Sciex 5500 QTrap mass spectrometer. This new ADE-OPP platform leverages the high speed and excellent accuracy of ADE liquid handling technology into sample delivery for mass spectrometry.

Matrix EffectsWe address the question: does decoupling of sample fluid delivery and ionization with ADE-OPP have an impact on ion suppression? It is known that ionization by sample fluid injection directly into gas phase may be impacted by ion suppression. Matrix effects were studied with 100 nanoM reserpine spiked into diluted digested plasma. For single droplet injection, no matrix effects were observed up to pure digested plasma with reserpine. With larger volumes, we observe onset of ion suppression at around 200 nL when using 10X dilution of plasma digest as matrix.

Open Port Probe Sampling InterfaceThe OPP sampling interface is a simple, versatile and self-cleaning system for rapidly introducing multiple types of unprocessed samples into a solvent flow stream for subsequent ionization and analysis by ESI mass spectrometry. This interface uses co-axial tubes arranged to deliver solvent to the sampling end of the device through the tubing annulus and then aspirate by the Venturi effect of the nebulizer gas through the center tube for delivery into the mass spectrometer ionization source.

Initial ResultsA precision solvent delivery pump allows us to explore the solvent flow rate effects on peak width and intensity. We tested a range of pump flow rates relative to the fixed aspiration rate, going from balanced flow rates with a small solvent vortex to unbalanced flows with a hanging droplet of solvent. The larger droplet volume of the hanging droplet leads to higher dilution and broader, lower intensity peaks. We observe that operation in Vortex mode with a single 2.5 nL droplet injection is desirable and Reserpine at 0.01 microM (25 amol) gives a 2 second peak at baseline. The flow rate stability of our pump provided good reproducibility and peak shape and allows us to generate calibration curves with excellent linearity. The continuous flow of solvent also mitigates carryover.

Conclusions• A new ADE-OPP platform leverages the speed and precision of ADE nanoliter droplets for

high-throughput MS.• Range of conditions explored to evaluate precision of the ADE-OPP combination with regard

to sample concentration, sample volume and sample matrix.• Flow control (key) and important for system stability and data precision.• Operation in Vortex Mode is desirable.• Good linearity with regards to volumes and concentrations.• Matrix effects explored to point where observed.

• Simple and robust

• Low pressure pump

• Up-right and upside-down

• Decouples sample delivery and ionization

• Continuous flow of solvent mitigates carryover

• No matrix effects observed for single drop

• Fast-response ~seconds per sample

Acoustic Droplet EjectionThe Labcyte Echo® ADE instrument is a robust platform that can eject 2.5 nL droplets from a fluid meniscus at high speed with excellent accuracy (< 10%) and precision (< 8% CV). The instrument can address a broad range of fluid classes utilizing Dynamic Fluid Analysis (DFA), a real-time auditing, sensing and calibration technology.

Acoustic Droplet Ejection into Open Port Probe (ADE-OPP)The Labcyte Echo® ADE instrument is customized to move the dispensing module external to commercial chassis for interface to Sciex 5500 QTrap MS.

Echo Transfer Volume Accuracy

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Echo Transfer Volume Precision

0%1%

2%3%4%5%

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CV

25 – 200%Triton X-100

0 – 60 %Glycerol

70 – 100%DMSO

0 – 100%FCS

25 – 200%Triton X-100

0 – 60 %Glycerol

70 – 100%DMSO

0 – 100%FCS

Open port probe

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Sample injected into vortex

Single droplet 2 second wide peak (baseline)

Vortex Suction upside down

Supercritical Critical Subcritical

Hanging droplet

Hanging dropletwith increasing size

Back to balanced

IncreasingMeOH flow

DecreasingMeOH flow

5 droplets (12.5 nL) injections in 1s (5Hz) 50 droplets (125 nL) injections in 1s (50Hz)

100 droplets (250 nL) injections in 1s (100Hz)10 droplets (25 nL) injections in 1s (10Hz)

25 droplets (62.5 nL) injections in 1s (25Hz) 200 droplets (500 nL) injections in 1s (200Hz)

y = 3350.9x + 15086R² = 0.9975

0.00E+00

1.00E+05

2.00E+05

3.00E+05

4.00E+05

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Number of Injected Droplets

10% β-gal 10% plasma

50% plasma 90% plasma Neat plasma

Balanced

1052-C