# US 10,770,277 B2 · System and method for the acoustic loading of an analytical instrument using a continuous flow sampling probe

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- Number: US 10,770,277 B2
- Document ID: US10770277B2
- Status: Granted
- Kind: B2
- Application number: 16/198,667
- Filed: 2018-11-21
- Published: 2019-05-23
- Granted: 2020-09-08
- Earliest claimed: 2017-11-22
- Assignees: Labcyte Inc.; DH Technologies Development Pte. Ltd.
- Applicants: Labcyte, Inc.; DH Technologies Development Pte. Ltd.
- Inventors: Sammy Datwani; Don W. Arnold; Lucien P. Ghislain; Chang Liu; Thomas Covey
- Web page: https://patentgrep.com/patent/US10770277B2

## CPC

- H01J 49/0454: {with means for vaporising using mechanical energy, e.g. by ultrasonic vibrations}
- B01L 3/0268: {using pulse dispensing or spraying, eg. inkjet type, piezo actuated ejection of droplets from capillaries}
- G01N 29/02: Analysing fluids (using acoustic emission techniques G01N29/14 {; constructional or flow details for analysing fluids G01N29/222; optoacoustic fluid cells G01N29/2425})
- G01N 29/222: {Constructional or flow details for analysing fluids (optoacoustic fluid cells G01N29/2425)}
- H01J 49/0031: {Step by step routines describing the use of the apparatus (H01J49/0081 takes precedence)}
- H01J 49/0404: {Capillaries used for transferring samples or ions (electrospray nozzles H01J49/167)}
- H01J 49/0445: {with means for introducing as a spray, a jet or an aerosol (electrospray ion sources H01J49/165)}
- H01J 49/045: {with means for using a nebulising gas, i.e. pneumatically assisted}
- H01J 49/165: {Electrospray ionisation}
- B01L 2400/0436: acoustic forces, e.g. surface acoustic waves [SAW]
- B01L 2400/0439: ultrasonic vibrations, vibrating piezo elements
- H01J: ELECTRIC DISCHARGE TUBES OR DISCHARGE LAMPS (spark-gaps H01T; electric arc lamps H05B31/00; particle accelerators H05H)
- B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

A system and method are provided for loading a sample into an analytical instrument using acoustic droplet ejection (“ADE”) in combination with a continuous flow sampling probe. An acoustic droplet ejector is used to eject small droplets of a fluid sample containing an analyte into the sampling tip of a continuous flow sampling probe, where the acoustically ejected droplet combines with a continuous, circulating flow stream of solvent within the flow probe. Fluid circulation within the probe transports the sample through a sample transport capillary to an outlet that directs the analyte away from the probe to an analytical instrument, e.g., a device that detects the presence, concentration quantity, and/or identity of the analyte. When the analytical instrument is a mass spectrometer or other type of device requiring the analyte to be in ionized form, the exiting droplets pass through an ionization region, e.g., an electrospray ion source, prior to entering the mass spectrometer or other analytical instrument. The method employs active flow control and enables real-time kinetic measurements.

## Claims

### Claim 1

An acoustic loading system for transporting an analyte in a fluid sample to an analytical instrument, comprising:
(a) a reservoir housing a fluid sample containing an analyte, the fluid sample having a fluid surface;
(b) an acoustic droplet ejector for generating acoustic radiation in a manner effective to eject a droplet of the fluid sample from the fluid surface; and
(c) a continuous flow sampling probe spaced apart from the fluid surface, comprising (i) a sampling tip for receiving the ejected droplet of the fluid sample, (ii) a solvent inlet for receiving a solvent from a solvent source, (iii) a solvent transport capillary for transporting the solvent from the solvent inlet to the sampling tip, where the ejected droplet combines with the solvent to form an analyte-solvent dilution, (iv) a sample outlet through which the analyte-solvent dilution is directed away from the sampling probe to an analytical instrument, and (v) a sample transport capillary for transporting the analyte-solvent dilution from the sampling tip to the sample outlet, wherein the sample transport capillary and the solvent transport capillary are in fluid communication at the sampling tip.

### Claim 2 (depends on 1)

The system of claim 1, wherein the continuous flow sampling probe comprises an outer capillary tube and an inner capillary tube co-axially disposed therein, the outer capillary tube and inner capillary tube defining the solvent transport capillary between the inner and outer capillary tubes and the inner capillary tube defining the sample transport capillary.

### Claim 3 (depends on 1)

The system of claim 1, wherein the acoustic droplet ejector comprises an acoustic radiation generator and a focusing means for focusing the acoustic radiation generated at a focal point near the surface of a fluid sample in the reservoir.

### Claim 4 (depends on 3)

The system of claim 3, wherein the acoustic droplet ejector is in acoustic coupling relationship with the reservoir.

### Claim 5 (depends on 1)

The system of claim 1, comprising a plurality of reservoirs each housing a fluid sample containing an analyte, wherein any one of the fluid samples may be the same or different as another of the fluid samples.

### Claim 6 (depends on 5)

The system of claim 5, further including a means for positioning the ejector in acoustic coupling relationship with respect to each of the reservoirs in succession.

### Claim 7 (depends on 5)

The system of claim 5, wherein the reservoirs are arranged in an array.

### Claim 8 (depends on 7)

The system of claim 7, wherein the reservoirs are contained within a substrate comprising an integrated multiple reservoir unit.

### Claim 9 (depends on 8)

The system of claim 8, further comprising a means for repositioning the reservoir unit relative to the acoustic ejector.

### Claim 10 (depends on 9)

The system of claim 9, wherein the fluid sample occupies a volume of no more than about 1 μL.

### Claim 11 (depends on 10)

The system of claim 10, wherein the fluid sample occupies a volume of about 10 pL to about 100 nL.

### Claim 12 (depends on 1)

The system of claim 1, wherein the acoustic droplet ejector is configured to eject a droplet having a volume of no more than about 3 nL.

### Claim 13 (depends on 12)

The system of claim 12, wherein the acoustic droplet ejector is configured to eject a droplet having a volume of no more than about 1 pL.

### Claim 14 (depends on 1)

The system of claim 1, further including a solvent pump operably connected to and in fluid communication with the solvent inlet for controlling solvent flow rate within the solvent transport capillary.

### Claim 15 (depends on 1)

The system of claim 1, further including a gas inlet through which a nebulizing gas flows from a gas source to the sample outlet, thereby aspirating the analyte-solvent dilution from the sample outlet.

### Claim 16 (depends on 15)

The system of claim 15, further including a gas pressure regulator operably connected to the gas inlet to control the nebulizing gas flow.

### Claim 17 (depends on 1)

The system of claim 1, further including an ionization source for ionizing analyte in the analyte-solvent dilution exiting the outlet.

### Claim 18 (depends on 17)

The system of claim 17, wherein the ionization source is an electrospray ion source.

### Claim 19 (depends on 18)

The system of claim 18, wherein the analytical instrument comprises a mass spectrometer.

### Claim 20 (depends on 1)

The system of claim 1, wherein the analytical instrument comprises a mass spectrometer.

### Claim 21 (depends on 1)

The system of claim 1, further including an adjuster adapted to move one of the inner capillary tube and the outer capillary tube longitudinally relative to the other.

### Claim 22

A method for transporting a fluid sample containing an analyte to an analytical instrument, the method comprising:
(a) acoustically coupling an acoustic droplet ejector that generates acoustic radiation to a reservoir containing the fluid sample having a fluid surface;
(b) activating the acoustic ejector to generate acoustic radiation toward the reservoir and into the fluid sample in a manner effective to eject a droplet of the fluid sample from the fluid surface into a sampling tip of a continuous flow sampling probe, where the ejected droplet combines with a circulating solvent within the flow probe to form an analyte-solvent dilution, said sampling probe spaced apart from the fluid surface to provide a gap between the fluid surface and the sampling tip; and
(c) transporting the received fluid sample droplet in a solvent through a sample transport capillary within the sampling probe to a sample outlet, where the analyte-solvent dilution is directed away from the sampling probe to an analytical instrument.

### Claim 23 (depends on 22)

The method of claim 22, wherein the continuous flow sampling probe comprises a solvent inlet for receiving the solvent from a solvent source and a solvent transport capillary for transporting the solvent from the solvent inlet to the sampling tip.

### Claim 24 (depends on 23)

The method of claim 23, wherein the sample transport capillary and the solvent transport capillary are in fluid communication at the sampling tip.

### Claim 25 (depends on 24)

The method of claim 24, wherein the continuous flow sampling probe comprises a gas inlet.

### Claim 26 (depends on 25)

The method of claim 25, wherein step (c) comprises flowing a nebulizing gas from a gas source through a gas inlet to the sample outlet to aspirate the analyte-solvent dilution at the sample outlet.

### Claim 27 (depends on 26)

The method of claim 26, further comprising controlling sample flow rate within the sample transport capillary with a gas pressure regulator operably connected to the gas inlet.

### Claim 28 (depends on 27)

The method of claim 27, further comprising controlling solvent flow rate within the solvent transport capillary with a solvent pump operably connected to and in fluid communication with the solvent inlet.

### Claim 29 (depends on 28)

The method of claim 28, further comprising adjusting the solvent flow rate relative to the sample flow rate to provide a desired flow pattern at the sampling tip where the solvent transport capillary and the sample transport capillary are in fluid communication.

### Claim 30 (depends on 29)

The method of claim 29, wherein the desired flow pattern is a vortex.

### Claim 31 (depends on 30)

The method of claim 30, wherein the vortex is a supercritical vortex.

### Claim 32 (depends on 22)

The method of claim 22, wherein the reservoir is one of a plurality of reservoirs each housing a fluid sample containing an analyte, wherein the fluid samples in the reservoirs may be the same or different.

### Claim 33 (depends on 32)

The method of claim 32, wherein the reservoirs are arranged in an array.

### Claim 34 (depends on 33)

The method of claim 33, wherein the reservoirs are contained within a substrate comprising an integrated multiple reservoir unit.

### Claim 35 (depends on 32)

The method of claim 32, further including, after ejecting the droplet, positioning the ejector in acoustic coupling relationship with respect to another reservoir and repeating steps (b) and (c).

### Claim 36 (depends on 22)

The method of claim 22, further comprising ionizing the analyte in the analyte-dilution exiting the sample outlet.

### Claim 37 (depends on 36)

The method of claim 36, wherein the analytical instrument comprises a mass spectrometer.

### Claim 38 (depends on 22)

The method of claim 22, wherein the analytical instrument comprises a mass spectrometer.

### Claim 39 (depends on 22)

The method of claim 22, wherein the analyte comprises a compound having a molecular weight in the range of about 100 daltons to about 100 kilodaltons.

### Claim 40 (depends on 39)

The method of claim 39, wherein the molecular weight is about 1 to about 100 kilodaltons.

### Claim 41 (depends on 40)

The method of claim 40, wherein the analyte is nonmetallic.

### Claim 42 (depends on 41)

The method of claim 41, wherein the analyte comprises an organic compound.

### Claim 43 (depends on 42)

The method of claim 42, wherein the organic compound is a biomolecule.

### Claim 44 (depends on 43)

The method of claim 43, wherein the biomolecule is peptidic.

### Claim 45 (depends on 22)

The method of claim 22, wherein the fluid sample occupies a volume of no more than about 10 μL.

### Claim 46 (depends on 45)

The method of claim 45, wherein the fluid sample occupies a volume of no more than about 1 μL.

### Claim 47 (depends on 46)

The method of claim 46, wherein the fluid sample occupies a volume of about 10 pL to about 100 nL.

### Claim 48 (depends on 22)

The method of claim 22, wherein the acoustically ejected droplet has a volume of no more than about 3 nL.

### Claim 49 (depends on 48)

The method of claim 48, wherein the acoustically ejected droplet has a volume of no more than about 1 pL.

### Claim 50 (depends on 48)

The method of claim 48, comprising rapidly repeating steps (a) and (b) to eject a plurality of ultra-monodisperse droplets into the sampling tip as one injection of the fluid sample.

### Claim 51 (depends on 50)

The method of claim 50, wherein steps (a) and (b) are repeated at a frequency of at least about 250 Hz.

### Claim 52 (depends on 22)

The method of claim 22, comprising rapidly repeating steps (a) and (b) to eject a plurality of ultra-monodisperse droplets into the sampling tip as one injection of the fluid sample.

### Claim 53 (depends on 22)

The method of claim 22, wherein the fluid sample comprises an analyte in a solvent.

### Claim 54 (depends on 22)

The method of claim 22, wherein the fluid sample comprises a biological fluid sample.

### Claim 55 (depends on 54)

The method of claim 54, wherein the biological fluid sample comprises tissue, tissue homogenate, cells, cell suspensions, cell extracts, whole blood, plasma, serum, saliva, sputum, nasal discharge, cerebrospinal fluid, interstitial fluid, lymph fluid, semen, vaginal fluid, or feces.