Apparatus and methods for controlling sonic treatment Number:7,521,023 from the United States Patent and Trademark Office (PTO) owispatent Apparatus and methods are disclosed for treating a sample by selectively controlling sonic energy and/or selectively controlling the location of the sample relative to the sonic energy.<H controlling, treatment, Apparatus, and, m, 7521023.html, Patent, pto, 7521023

Title: Apparatus and methods for controlling sonic treatment

Abstract: Apparatus and methods are disclosed for treating a sample by selectively controlling sonic energy and/or selectively controlling the location of the sample relative to the sonic energy.

Patent Number: 7,521,023 Issued on 04/21/2009 to Laugharn, Jr., et al.

Inventors: Laugharn, Jr.; James A. (Winchester, MA), Garrison; Brevard S. (Reading, MA)
Assignee: Covaris, Inc. (Woburn, MA)

Appl. No.: 10/777,014

Filed: February 11, 2004

Related U.S. Patent Documents


Application Number	Filing Date	Patent Number	Issue Date
09830473		6719449
PCT/US99/25274	Oct., 1999
60148279	Aug., 1999
60143440	Jul., 1999
60119500	Feb., 1999
60110460	Dec., 1998
60105933	Oct., 1998

Current U.S. Class: 422/128 ; 366/127

Current International Class: B06B 1/00 (20060101); B01F 11/02 (20060101)

Field of Search: 366/132,151.1,152.2,127,116 422/127,128,20

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Primary Examiner: Soohoo; Tony G
Attorney, Agent or Firm: Wolf, Greenfield & Sacks, P.C.

Parent Case Text

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No. 09/830,473, filed Apr. 27, 2001 (U.S. Pat. No. 6,719,449), which is the U.S. national phase of International (PCT) Patent Application Ser. No. PCT/US99/25274, filed on Oct. 28, 1999, and published in English on May 4, 2000, which claims the benefit of and priority to U.S. provisional application Ser. Nos. 60/105,933, filed Oct. 28, 1998; No. 60/110,460, filed Dec. 1, 1998; No. 60/119,500, filed Feb. 10, 1999; No. 60/143,440, filed Jul. 13, 1999; and No. 60/148,279, filed Aug. 11, 1999, the entirety of each of the disclosures of these applications being incorporated by reference herein.

Claims

What is claimed is:

1. An apparatus for processing at least one sample, comprising: (a) vessel for holding the at least one sample and including at least one inlet for flowing the at least one sample in a liquid medium into the vessel and at least one outlet for flowing the at least one sample in the liquid medium out of the vessel; and (b) an acoustic energy source spaced from and exterior to the vessel for providing at least one focused acoustic field having a frequency of between about 100 kilohertz and about 100 megahertz and having a focal zone having a width of less than about 2 centimeters to the at least one sample while the at least one sample is in the vessel, wherein at least a portion of acoustic energy from the acoustic energy source propagates exterior to the vessel.

2. The apparatus of claim 1, wherein the at least one focused acoustic field has a focal zone smaller than the vessel.

3. The apparatus of claim 1, wherein the at least one focused acoustic field has a focal zone larger than the vessel.

4. The apparatus of claim 1, wherein the at least one focused acoustic field includes a plurality of focused acoustic fields and the acoustic energy source includes a plurality of acoustic transducers for providing the plurality of the focused acoustic fields to the at least one sample.

5. The apparatus of claim 1, including a positioning system for positioning the at least one sample and the at least one focused acoustic field relative to each other.

6. The apparatus of claim 1, including a processor for controlling a positioning system to stop sample movement relative to the acoustic energy source to facilitate treating of the at least one sample.

7. The apparatus of claim 1, including a processor for controlling a positioning system to dither a relative position of the at least one sample and the focal zone.

8. The apparatus of claim 1, wherein the at least one sample includes at least one of a mineral or a biological material.

9. The apparatus of claim 1, wherein the at least one sample is suspended in the liquid medium.

10. The apparatus of claim 9, wherein the liquid medium includes a solvent.

11. The apparatus of claim 1, wherein the at least one sample includes a first molecule and a second molecule, and the second molecule is different from the first molecule.

12. The apparatus of claim 11, wherein the at least one sample includes a first nucleic acid molecule and the second molecule includes a second nucleic acid molecule.

13. The apparatus of claim 12, wherein the first nucleic acid molecule is a primer and the second nucleic acid molecule is a substrate molecule.

14. The apparatus of claim 1, wherein the at least one sample includes an antibody and a molecule to which the antibody binds.

15. The apparatus of claim 1, wherein the at least one sample includes a substrate and a ligand.

16. The apparatus of claim 1, wherein the at least one sample includes at least one of an antibody and a receptor and further comprising a support surface for immobilizing the at least one of the antibody and the receptor.

17. The apparatus of claim 1, wherein the at least one sample is heated in response to the at least one focused acoustic field.

18. The apparatus of claim 1, wherein a portion of the at least one sample is disrupted in response to the at least one focused acoustic field.

19. The apparatus of claim 1, wherein permeability of the at least one sample is increased in response to the at least one focused acoustic field.

20. The apparatus of claim 1, wherein a reaction within the at least one sample is enhanced in response to the at least one focused acoustic field.

21. The apparatus of claim 1, wherein extra-cellular membranes of the at least one sample are disrupted in response to the at least one focused acoustic field.

22. The apparatus of claim 1, wherein a barrier function of a structure in the at least one sample is lessened in response to the at least one focused acoustic field.

23. The apparatus of claim 1, including a processor for controlling the acoustic energy source to be on during a treat interval and off during a dead interval.

24. The apparatus of claim 23, wherein the processor controls a frequency of operation of the acoustic energy source.

25. The apparatus of claim 23, wherein the processor controls a duty cycle of operation of the acoustic energy source.

26. The apparatus of claim 1, further comprising a system for transferring the vessel into or out of a treatment apparatus.

27. The apparatus of claim 1, including a processor for controlling flow of the liquid medium into and out of the vessel to control exposure of the at least one sample to the at least one focused acoustic field.

28. The apparatus of claim 1, including a processor for controlling the acoustic energy source to control exposure of the at least one sample to the at least one focused acoustic field.

29. The apparatus of claim 1, including a processor for varying a frequency of the acoustic energy source to control exposure of the at least one sample to the at least one focused acoustic field.

30. The apparatus of claim 1, including a feedback system having a sensor for providing feedback information relevant to the at least one sample.

31. The apparatus of claim 30, including a processor for determining a state of treatment of the at least one sample based, at least in part, on the feedback information.

32. The apparatus of claim 31, wherein the processor controls flow of the at least one sample and the liquid medium based, at least in part, on determination of the state of treatment.

33. The apparatus of claim 31, wherein the processor controls a transducer based, at least in part, on determination of the state of treatment.

34. The apparatus of claim 30, wherein the sensor includes an acoustic transducer for detecting acoustic emissions from the liquid medium.

35. The apparatus of claim 30, wherein the sensor includes an acoustic transducer for detecting acoustic reflections from the at least one sample.

36. The apparatus of claim 30, wherein the sensor includes a temperature sensor and the feedback information includes temperature information.

37. The apparatus of claim 30, wherein the sensor includes optical detection and the feedback information includes spectral information.

38. The apparatus of claim 37, wherein the spectral information includes at least one of spectral excitation, absorption, fluorescence, and emission of the at least one sample.

39. The apparatus of claim 1, further comprising a coupling medium for coupling the focused acoustic field to the at least one sample, wherein said coupling medium does not contact the at least one sample.

40. The apparatus of claim 1 wherein the vessel is a conduit.

41. The apparatus of claim 40, wherein the at least one sample is held within a separate container included within the conduit.

42. The apparatus of claim 1, wherein the vessel comprises a plurality of containers each capable of holding at least one sample.

43. The apparatus of claim 42, wherein the vessel comprises at least one of a microtiter plate, a blister pack, and an array of polymeric bubbles.

44. The apparatus of claim 1, wherein the focused acoustic field has a focal zone with a width of less than about 1 cm.

45. The apparatus of claim 1, wherein the focused acoustic field has a focal zone with a width of less than about 1 mm.

46. The apparatus of claim 1, wherein the width corresponds to a diameter of the focal zone.

47. The apparatus of claim 1, wherein the acoustic energy source for providing the at least one focused acoustic field is modulated to produce multiple foci.

48. The apparatus of claim 1, wherein the acoustic energy source for providing the at least one focused acoustic field generates a cigar-shaped focal zone.

49. The apparatus of claim 1, wherein the acoustic energy source for providing the at least one focused acoustic field generates an ellipsoidal shaped focal zone.

50. The apparatus of claim 1, wherein the at least one sample is a fluid.

51. The apparatus of claim 1, wherein the at least one sample and the liquid medium are substantially similar.

52. The apparatus of claim 1, wherein the at least one focused acoustic field sterilizes the at least one sample.

53. The apparatus of claim 1, wherein the at least one focused acoustic field aids the at least one sample to flow toward the at least one outlet.

54. The apparatus of claim 1, wherein the vessel has a width of at least about 10 mm.

55. The apparatus of claim 1, wherein the vessel comprises a treatment chamber, the treatment chamber having substantially vertical walls.

56. The apparatus of claim 1, wherein the vessel comprises a treatment chamber, the treatment chamber having a portion that is substantially conical in shape.

57. The apparatus of claim 1, wherein the vessel comprises a treatment chamber, the treatment chamber having a portion that is substantially hemispherical in shape.

58. The apparatus of claim 1, wherein flow of the at least one sample in a liquid medium comprises laminar flow.

59. The apparatus of claim 1, wherein flow of the at least one sample in a liquid medium comprises turbulent flow.

60. The apparatus of claim 1, wherein the at least one acoustic field propagates with sufficient energy to induce cavitation in the liquid medium.

61. The apparatus of claim 1, further comprising: a container for holding a liquid coupling medium through which acoustic energy from the acoustic energy source travels to the vessel and the at least one sample, the liquid coupling medium contacting the vessel but not contacting the at least one sample.

62. The apparatus of claim 61, wherein the acoustic energy source is located in the container so as to be in contact with the liquid coupling medium when the liquid coupling medium is held by the container.

Description

TECHNICAL FIELD

The present invention generally relates to the field of controlled sonic energy emittingdevices for treating material, particularly biological material.

BACKGROUND OF THE INVENTION

Ultrasonics have been utilized for many years for a variety of diagnostic, therapeutic, and research purposes. The acoustic physics of ultrasonics is well understood; however, the biophysical, chemical, and mechanical effects are generally only empirically understood. Some uses of sonic or acoustic energy in materials processing include "sonication," an unrefined process of mechanical disruption involving the direct immersion of an unfocused ultrasound source emitting energy in the kilohertz ("kHz") range into a fluid suspension of the material being treated. Accordingly, the sonic energy often does not reach a target in an effective dose because the energy is scattered, absorbed, and/or not properly aligned with the target. There are also specific clinical examples of the utilization of therapeutic ultrasound (e.g., lithotripsy) and of diagnostic ultrasound (e.g., fetal imaging). However, ultrasonics have heretofore not been controlled to provide an automated, broad range, precise materials processing or reaction control mechanism.

SUMMARY OF THE INVENTION

The present invention relates to apparatus and methods for selectively exposing a sample to sonic energy, such that the sample is exposed to produce a desired result such as, but without limitation, heating the sample, cooling the sample, fluidizing the sample, mixing the sample, stirring the sample, disrupting the sample, permeabilizing a component of the sample, enhancing a reaction in the sample, and sterilizing the sample. For example, altering the permeability or accessibility of a material, especially labile biological materials, in a controlled manner can allow for manipulation of the material while preserving the viability and/or biological activity of the material. In another example, mixing materials or modulating transport of a component into or out of materials, in a reproducible, uniform, automated manner, can be beneficial. According to one embodiment of the system, sample processing control includes a feedback loop for regulating at least one of sonic energy location, pulse pattern, pulse intensity, and absorbed dose of the ultrasound. The system can be automated. In one embodiment, the ultrasonic energy is in the megahertz (MHz) frequency range, in contrast to classical sonic processing which typically employs ultrasonic energy in the kilohertz (kHz) frequency range.

When ultrasonic energy interacts with a complex biological or chemical system, the acoustic field often becomes distorted, reflected, and defocused. The net effect is that energy distribution becomes non-uniform and/or defocused compared to the input. Non-uniform reaction conditions can limit reaction applications to non-critical processes, such as bulk fluid treatment where temperature gradients within a sample are inconsequential. However, some of the non-uniform aspects are highly deleterious to samples, such as extreme temperature gradients that damage sample integrity. For example, in some instances, the high temperature would irreversibly denature target proteins. As a consequence, many potential applications of ultrasound, especially biological applications, are limited to specific, highly specialized applications, such as lithotripsy and diagnostic imaging, because of the potentially undesirable and uncontrollable aspects of ultrasound in complex systems.

Typically, when ultrasound is applied to a bulk biological sample solution, such as for the extraction of intracellular constituents from tissue, the treatment causes a complex, heterogeneous, mixture of sub-events that vary during the course of a treatment dose. In other words, the ultrasonic energy may be partitioned between various states. For example, the energy may directly treat a sample or the energy may spatially displace a target moiety and shift the target out of the optimal energy zone. Additionally or alternatively, the energy may result in interference that reflects the acoustic energy. For example, a "bubble shield" occurs when a wave front of sonic energy creates cavitation bubbles that persist until the next wave front arrives, such that the energy of the second wave front is at least partially blocked and/or reflected by the bubbles. Still further, larger particles in the sample may move to low energy nodes, thereby leaving the smaller particles in the sample with more dwell-time in the high energy nodes. In addition, the sample viscosity, temperature, and uniformity may vary during the ultrasonic process, resulting in gradients of these parameters during processing. Accordingly, current processes are generally random and non-uniform, especially when applied to in vitro applications, such as membrane permeabilization, hindering the use of ultrasound in high throughput applications where treatment standardization from one sample to the next is required.

Processing samples containing labile material, in particular biological material, is still largely a manual process, and poorly adapted to high-throughput sample processing required for applications such as pharmaceutical and agricultural genomics. For example, except for isolated or exposed cells, the insertion of a nucleic acid into a sample, for temporary or permanent transformation, is still substantially manual. Most transformation techniques have been developed for a small subset of materials, which typically have only a single plasma membrane separating their interior from the environment. These membranes may be permeabilized using detergents, salts, osmotic shock, or simple freeze-thawing. Thus, materials such as viruses, cultured cells, and bacteria and protists, such as yeast, which have been treated to prevent the formation of cell walls, can be transfected by any of a number of standard methods. For example, transfection can be undertaken with vectors including viruses that bind to plasma membranes for direct transport, and can be undertaken in a direct transfection with "naked" DNA that is often coated with cationic lipids or polymers or that is in the presence of chemical or biochemical membrane permeabilizing agents.

Moreover, many biological materials of interest have supporting structures, and are significantly harder to permeabilize or otherwise to access the plasma membrane with macromolecular agents or viruses. The supporting structures range from simple cell walls, as in yeast, to complex protein and glycoprotein structures, as in animal tissue, to tenacious and only slowly degradable polysaccharide structures, as in plants and insects, to physically durable mineralized supports, as in diatoms and bone. In all of these "hard" materials, physical disruption of the supporting matrices is required typically to precede or accompany transfection or other nucleic acid insertion to allow reliable introduction of extracellular components.

Sonication has been used to break up difficult materials such as plant tissue. Sonication, typically implemented by vibration of a probe at frequencies of 10,000 Hz or higher, creates shearing forces within a liquid sample. However, the resultant shear is not readily controlled, so that when sufficient energy is applied to disrupt a supporting matrix, the shear will also tend to destroy fragile intracellular structures. Indeed, sonication is routinely used to randomly shear DNA in solution into small fragments. Such fragmentation limits the usefulness of these techniques for many purposes, and particularly for transfection, which requires a viable cell to be successful.

The present invention addresses these problems and provides apparatus and methods for the non-contact treatment of samples with ultrasonic energy, using a focused beam of energy. The frequency of the beam can be variable and can be in the range of about 100 kHz to 100 MHz, more preferably 500 kHz to 10 MHz. For example, the present invention can treat samples with ultrasonic energy while controlling the temperature of the sample, by use of computer-generated complex wavetrains, which may further be controlled by the use of feedback from a sensor. The acoustic output signal, or wavetrain, can vary in any or all of frequency, intensity, duty cycle, burst pattern, and pulse shape. In another example, the present invention can treat samples with ultrasonic energy when the samples are in an array, and individual samples in the array may be treated differentially or identically. Moreover, this treatment can be undertaken automatically under computer control. In another example, the present invention can treat samples with ultrasonic energy in a uniform way over the entire sample, by the relative movement of the sample and the focus of the beam, in any or all of two or three dimensions.

The apparatus and methods of the present invention can be controlled by a computer program. In one embodiment, the sequence of actions taken by the computer is predetermined. Such embodiments can be useful in high-speed, high-volume processing. In another embodiment, the processes are enhanced with a program that uses feedback control to modify or determine the actions thereof, using techniques including algorithmic processing of input, the use of lookup tables, and similar integration devices and processes.

A feedback control mechanism, in connection with any of accuracy, reproducibility, speed of processing, control of temperature, provision of uniformity of exposure to sonic pulses, sensing of degree of completion of processing, monitoring of cavitation, and control of beam properties (including intensity, frequency, degree of focusing, wavetrain pattern, and position), can enhance certain embodiments of the present invention. A variety of sensors or sensed properties may be appropriate for providing input for feedback control. These properties can include sensing of temperature of the sample; sonic beam intensity; pressure; bath properties including temperature, salinity, and polarity; sample position; and optical or visual properties of the samples. These optical properties may include apparent color, emission, absorption, fluorescence, phosphorescence, scattering, particle size, laser/Doppler fluid and particle velocities, and effective viscosity. Sample integrity or comminution can be sensed with a pattern analysis of an optical signal. Any sensed property or combination thereof can serve as input into a control system. The feedback can be used to control any output of the system, for example beam properties, sample position, and treatment duration.

The samples can be treated in any convenient vessel or container. Vessels can be sealed for the duration of the treatment to prevent contamination of the sample or of the environment. Arrays of vessels can be used for processing large numbers of samples. These arrays can be arranged in one or more high throughput configurations. Examples include microtiter plates, typically with a temporary sealing layer to close the wells, blister packs, similar to those used to package pharmaceuticals such as pills and capsules, and arrays of polymeric bubbles, similar to bubble wrap, preferably with a similar spacing to typical microtiter wells. The latter are described in more detail below.

The treatment, which may be performed or enhanced by use of ultrasonic wavetrains, include any unit operation which is susceptible to being implemented or is enhanced by sonic waves or pulses. In particular, these results include lysing, extracting, permeabilizing, stirring or mixing, comminuting, heating, fluidizing, sterilizing, catalyzing, and selectively degrading. Sonic waves may also enhance filtration, fluid flow in conduits, and fluidization of suspensions. Processes of the invention may be synthetic, analytic, or simply facilitative of other processes such as stirring.

Any sample is potentially suitable for processing by the techniques and apparatuses of the invention. For example, any material that includes biological organisms or material derived therefrom is suitable. Many chemicals can be processed more efficiently, particularly in small-scale or combinatorial reactions or assays, with the processes of the invention, including remote, non-contact mixing or stirring. Physical objects, such as mineral samples and particulates including sands and clays, also can be treated with the present invention.

According to the present invention, several aspects of the invention can enhance the reproducibility and/or effectiveness of particular treatments using ultrasonic energy in in vitro applications, where reproducibility, uniformity, and precise control are desired. These aspects include the use of feedback, precise focusing of the ultrasonic energy, monitoring and regulating of the acoustic waveform (including frequency, amplitude, duty cycle, and cycles per burst), positioning of the reaction vessel relative to the ultrasonic energy so that the sample is uniformly treated, controlling movement of the sample relative to the focus of ultrasonic energy during a processing step, and/or controlling the temperature of the sample being treated, either by the ultrasonic energy parameters or through the use of temperature control devices such as a water bath. A treatment protocol can be optimized, using one or a combination of the above variables, to maximize, for example, shearing, extraction, permeabilization, comminution, stirring, or other process steps, while minimizing undesirable thermal effects.

In one embodiment of the invention, high intensity ultrasonic energy is focused on a reaction vessel, and "real time" feedback relating to one or more process variables is used to control the process. In another embodiment, the process is automated and is used in a high throughput system, such as a 96-well plate, or a continuous flowing stream of material to be treated, optionally segmented.

Minimization of unwanted interference with the pattern of applied ultrasonic energy is another feature of the invention. For example, ultrasonic energy applied to a sample in a reaction vessel has the potential to directly interact with the target sample, or to reflect from bubbles or other effects from a previous cycle of ultrasound application and not interact with the target, or to miss the target because of spatial separation or mismatch. Minimization of interference is especially beneficial for remote, automated, sterile processing of small amounts of target material, for example, 10 mg of a biopsy tissue. By minimizing the reflections and optimizing spatial positioning, the ultrasonic energy is more efficiently utilized and controlled. The process can be standardized to obtain reproducibility by presetting conditions such as waveform and positioning, by a feedback signal and feedback-based control to maintain preset performance target parameters, or by a combination of these methods.

In certain embodiments, the processing system can include a high intensity transducer that produces acoustic energy when driven by an electrical or optical energy input; a device or system for controlling excitation of the transducer, such as an arbitrary waveform generator, an RF amplifier, and a matching network for controlling parameters such as time, intensity, and duty cycle of the ultrasonic energy; a positioning system such as a 2-dimensional (x, y) or a 3-dimensional (x, y, z) positioning system that can be computer controlled to allow automation and the implementation of feedback from monitoring; a temperature sensor; a device for controlling temperature; one or more reaction vessels; and a sensor for detecting, for example, optical, radiative, and/or acoustic signatures.

Vessels containing the samples can be sealed during the processing, and hence can be sterile throughout, or after, the procedure. Moreover, the use of focused ultrasound allows the samples in the vessels to be processed, including processing by stirring, without contacting the samples, even when the vessels are not sealed.

The processes have a variety of applications, including, but without limitation, extraction, permeabilization, mixing, comminuting, sterilization, flow control, and reacting. For example, mixing in a vessel can be achieved with temperature fluctuations controlled to within about plus or minus one degree Celsius. More precise control is possible, if required. In another example, labile biological materials can be extracted from plant materials without loss of activity or the use of harsh solvents. In other applications, complex cells can be permeabilized and molecules such as nucleotide molecules can be introduced into the cells using the process of the invention. Other applications include modulating binding reactions that are useful in separations, biological assays, and hybridization reactions.

One aspect of the invention includes an apparatus for processing a sample using sonic energy. The apparatus includes a sonic energy source for emitting sonic energy; a holder for the sample, the sample movable relative to the emitted sonic energy; and a processor for controlling the sonic energy source and location of the sample according to a predetermined methodology, such that the sample is selectively exposed to sonic energy to produce a desired result. The desired result can be heating the sample, cooling the sample, fluidizing the sample, mixing the sample, stirring the sample, disrupting the sample, increasing permeability of a component of the sample, enhancing a reaction within the sample, and/or sterilizing the sample. Also, the desired result can be an in vitro or an ex vivo treatment.

This aspect and other aspects of the invention can include any or all of the following features. The apparatus can further include a feedback system connected to the processor for monitoring at least one condition to which the sample is subjected during processing, such that the processor controls at least one of the sonic energy source and the location of the sample in response to the at least one condition. The feedback system can include a sensor for monitoring the at least one condition. The apparatus can further include a temperature control unit for controlling temperature of the sample, and the processor can control the temperature control unit. The apparatus can further include a pressure control unit for controlling pressure to which the sample is exposed, and the processor controls the pressure control unit. The sonic energy source can include a transducer. The transducer can focus the sonic energy and can include at least one piezoelectric element, an array of piezoelectric elements, an electrohydraulic element, a magnetostrictive element, an electromagnetic transducer, a chemical explosive element, and/or a laser-activated element. A piezoelectric element can include a spherical transmitting surface oriented such that the focal axis is oriented vertically or in any other predetermined direction. The holder can support a sample container for containing the sample. The sample container can be a membrane pouch, thermopolymer well, polymeric pouch, hydrophobic membrane, microtiter plate, microtiter well, test tube, centrifuge tube, microfuge tube, ampoule, capsule, bottle, beaker, flask, and/or capillary tube. The sample container can form multiple compartments and can include a rupturable membrane for transferring a fraction of the sample away from the holder. The apparatus can further include a device for moving the sample from a first location to a second location, such as a stepper motor. The apparatus can also include an acoustically transparent material disposed between the sonic energy source and the holder. The sample can flow through a conduit. The sonic energy source can generate sonic energy at two or more different frequencies, optionally in the form of a serial wavetrain. The wavetrain can include a first wave component and a different second wave component. Alternatively or additionally, the wavetrain can include about 1000 cycles per burst at about a 10% duty cycle at about a 500 mV amplitude.

Another aspect of the invention relates to a method for processing a sample with sonic energy. The method includes the steps of exposing the sample to sonic energy and controlling at least one of the sonic energy and location of the sample relative to the sonic energy according to a predetermined methodology, such that the sample is selectively exposed to sonic energy to produce a desired result. The desired result can be heating the sample, cooling the sample, fluidizing the sample, mixing the sample, stirring the sample, disrupting the sample, increasing permeability of a component of the sample, enhancing a reaction within the sample, and/or sterilizing the sample. Also, the desired result can be an in vitro or an ex vivo treatment. This aspect or any of the other aspects of the invention can include any or all of the following features. The method can further include the steps of sensing at least one condition to which the sample is subjected during processing and altering at least one of the sonic energy and the location of the sample in response to the sensed condition. During the sensing step, the sensed condition can be temperature, pressure, an optical property, an altered chemical, an acoustic signal, and/or a mechanical occurrence. During the altering step, the characteristic of the sonic energy that is altered can be waveform, duration of application, intensity, and/or duty cycle. The method can further include the step of controlling temperature of the sample and can further include the step of controlling pressure to which the sample is exposed. During the step of exposing the sample to sonic energy, the sonic energy can be generated by spark discharges across a gap, laser pulses, piezoelectric pulses, electromagnetic shock waves, electrohydraulic shock waves, electrical discharges into a liquid, and/or chemical explosives. The sonic energy can be focused on the sample. The sample can contain a cell, and the method can further comprise the step of introducing a material into the cell. The material can be a polymer, an amino acid monomer, an amino acid chain, a protein, an enzyme, a nucleic acid monomer, a nucleic acid chain, a saccharide, a polysaccharide, an organic molecule, an inorganic molecule, a vector, a plasmid, and/or a virus. The method can further include the step of extracting a component of the sample. During the controlling step, at least one characteristic of the sonic energy is controlled, that characteristic being waveform, duration of application, intensity, or duty cycle. The method can further include the step of the sample flowing through a conduit. The sonic energy can include at least two different frequencies, optionally in the form of a wavetrain. The wavetrain can include a first wave component and a different second wave component. Alternatively or additionally, the wavetrain can include about 1000 cycles per burst at about a 10% duty cycle at about a 500 mV amplitude.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention, in accordance with preferred and exemplary embodiments, together with further advantages thereof, is more particularly described in the following detailed description, taken in conjunction with the accompanying drawings.

In the drawings, like reference characters generally refer to the same parts throughout the different views. Also, the drawings are not necessarily to scale, emphasis instead generally being placed upon illustrating principles of the invention.

FIG. 1 is a schematic illustration of one embodiment of the apparatus according to the invention;

FIG. 2 is a schematic illustration of one example of sonic energy control showing sine waves at a variable amplitude and frequency;

FIG. 3 is a schematic illustration of one example of an intra-sample positioning (dithering) profile showing height, height step, and radius;

FIG. 4A is a schematic illustration of a vertical-sided treatment vessel;

FIG. 4B is a schematic illustration of a conical treatment vessel;

FIG. 4C is a schematic illustration of a curved treatment vessel;

FIGS. 5A-5C are schematic illustrations of several embodiments of a treatment vessel with a combination of an upper and lower member and samples in the vessels prior to treatment;

FIG. 6A is a schematic illustration of a treatment vessel positioned over a collection container prior to transferring the contents of the vessel to the container;

FIG. 6B is a schematic illustration of a treatment vessel positioned over a collection container after transferring some of the contents of the vessel to the container;

FIG. 7 is a schematic illustration of an in-line fluid treatment method in accordance with an alternative embodiment of the invention;

FIG. 8 is a graph depicting change in sample temperature as a function of duty cycle at 500 mV and 750 mV, in one embodiment of the invention;

FIG. 9 is a schematic illustration of an embodiment of the invention with a microtiter plate containing samples, such that one of the wells of the microtiter plate is positioned at the focus point of sonic energy;

FIG. 10 describes certain features and specifications related to performance, consumables, procedure for treatment, and mechanical components of a system according to certain embodiments of the invention;

FIG. 11 describes certain features and specifications related to instrument control, user interface, electrical, and associated equipment of a system according to certain embodiments of the invention;

FIG. 12 describes certain characteristics and functionality of operating software related to general functions, display functions, sonic energy control, and target/source positioning of a system according to certain embodiments of the invention; and

FIG. 13 describes certain additional characteristics and functionality of operating software related to target/source positioning and temperature control of a system according to certain embodiments of the invention.

DETAILED DESCRIPTION OF THE INVENTION

"Sonic energy" as used herein is intended to encompass such terms as acoustic energy, acoustic waves, acoustic pulses, ultrasonic energy, ultrasonic waves, ultrasound, shock waves, sound energy, sound waves, sonic pulses, pulses, waves, or any other grammatical form of these terms, as well as any other type of energy that has similar characteristics to sonic energy. "Focal zone" or "focal point" as used herein means an area where sonic energy converges and/or impinges on a target, although that area of convergence is not necessarily a single focused point. As used herein, the terms "microplate," "microtiter plate," "microwell plate," and other grammatical forms of these terms can mean a plate that includes one or more wells into which samples may be deposited. As used herein, "nonlinear acoustics" can mean lack of proportionality between input and output. For example, in our application, as the amplitude applied to the transducer increases, the sinusoidal output loses proportionality such that eventually the peak positive pressure increases at a higher rate than the peak negative pressure. Also, water becomes nonlinear at high intensities, and in a converging acoustic field, the waves become more disturbed as the intensity increases toward the focal point. Nonlinear acoustic properties of tissue can be useful in diagnostic and therapeutic applications. As used herein, "acoustic streaming" can mean generation of fluid flow by acoust