Search All Patents:

---

Register or Login To Download This Patent As A PDF

---

United States Patent Application	20120024867
Kind Code	A1
Laugharn, JR.; James A.	February 2, 2012

---

---

---

Claims

---

1. A method for acoustic treatment of a sample contained in a vessel, comprising: providing a vessel containing a liquid sample, the vessel having a wall in contact with the liquid sample, the wall including a heat exchanger on an inner surface that is in contact with the liquid sample; applying acoustic energy from an acoustic energy source to the liquid sample to cause movement of portions of the liquid sample near the vessel wall; and using the heat exchanger on the inner surface of the vessel wall to interact with moving portions of the liquid sample and disrupt a boundary layer of the liquid sample at the vessel wall, disruption of the boundary layer enhancing heat transfer between the vessel wall and the liquid sample.

2. The method of claim 1, further comprising: simultaneous with disrupting the boundary layer, applying acoustic energy from the acoustic energy source to the vessel wall to heat the vessel wall and increase the vessel wall's temperature above a temperature of the liquid sample.

3. The method of claim 2, wherein heating the vessel wall causes heat transfer from the vessel wall to the liquid sample to raise the temperature of the liquid sample above a temperature of a coupling medium in contact with an exterior of the vessel.

4. The method of claim 1, wherein a temperature of the vessel wall is below a temperature of the liquid sample, and disrupting the boundary layer causes heat transfer from the liquid sample to the vessel wall so as to lower a temperature of the liquid sample.

5. The method of claim 1, wherein the heat exchanger includes a plurality of raised areas and/or grooves on the vessel wall.

6. The method of claim 1, wherein the heat exchanger extends around an entire internal periphery of the vessel wall and extends along at least a portion of a length of the vessel.

7. The method of claim 1, wherein the heat exchanger includes features that are molded integrally with the vessel wall.

8. The method of claim 1, wherein the vessel wall includes a heat exchanger on an outer surface to interact with an environment outside of the vessel.

9. The method of claim 1, simultaneous with disrupting the boundary layer, applying acoustic energy from the acoustic energy source to the vessel wall to heat the vessel wall and heat the liquid sample at a rate of at least about 25 degrees C. per ml per minute.

10. A method for acoustic treatment of a sample contained in a vessel, comprising: providing a vessel containing a liquid sample, the vessel having a wall in contact with the liquid sample, the wall including a heat exchanger on an outer surface; providing a coupling medium in contact with the heat exchanger of the vessel, the coupling medium having a temperature that is different from a temperature of the liquid sample; and disrupting a boundary layer between the liquid sample and the wall by transmitting acoustic energy through the coupling medium and to the vessel and liquid sample to cause movement of portions of the liquid sample, disruption of the boundary layer enhancing heat transfer between the liquid sample and the vessel wall.

11. The method of claim 10, wherein the heat exchanger includes a plurality of raised areas and/or grooves on the outer surface of the vessel wall.

12. The method of claim 10, further comprising: simultaneous with disrupting the boundary layer, applying acoustic energy from the acoustic energy source to the vessel wall to heat the vessel wall and increase the vessel wall's temperature above a temperature of the liquid sample.

13. The method of claim 12, wherein heating the vessel wall causes heat transfer from the vessel wall to the liquid sample to raise the temperature of the liquid sample above the temperature of the coupling medium.

14. The method of claim 10, wherein a temperature of the vessel wall is below a temperature of the liquid sample, and disrupting the boundary layer causes heat transfer from the liquid sample to the vessel wall so as to lower a temperature of the liquid sample.

15. The method of claim 10, wherein the vessel is one of a plurality of vessels in a multi-well plate.

16. The method of claim 10, wherein the heat exchanger extends around an entire external periphery of the vessel wall and extends along at least a portion of a length of the vessel.

17. The method of claim 10, wherein the heat exchanger includes features that are molded integrally with the vessel wall.

18. The method of claim 10, wherein the vessel wall includes a heat exchanger on an inner surface to interact with the liquid sample.

19. The method of claim 10, simultaneous with disrupting the boundary layer, applying acoustic energy from the acoustic energy source to the vessel wall to heat the vessel wall and heat the liquid sample at a rate of at least about 25 degrees C. per ml per minute.

20. The method of claim 10, wherein the temperature of the coupling medium is lower than a temperature of the liquid sample, and the step of disrupting causes a temperature of the liquid sample to be reduced.

21. A method for acoustic treatment of a sample contained in a vessel, comprising: providing a vessel containing a liquid sample, the vessel having a wall with an inner surface in contact with the liquid sample; providing a coupling medium in contact with an outer surface of the vessel; applying acoustic energy from an acoustic energy source through the coupling medium to the vessel wall to heat the vessel wall and increase the vessel wall's temperature above a temperature of the liquid sample; and simultaneous with applying acoustic energy to heat the vessel wall, applying acoustic energy from the acoustic energy source to the liquid sample to disrupt a boundary layer of the liquid sample at the vessel wall so as to enhance heat transfer from the vessel wall to the liquid sample and to raise the temperature of the liquid sample above a temperature of the coupling medium, heating of the liquid sample being performed at a rate of at least about 25 degrees C. per ml per minute.

22. The method of claim 21, wherein the coupling medium that couples acoustic energy from the acoustic energy source to the vessel wall is a liquid.

23. The method of claim 21, wherein the coupling medium is at a temperature that is lower than a temperature of the liquid sample.

24. The method of claim 21, further comprising: subsequent to heating the liquid sample to a temperature greater than the temperature of the coupling medium, stopping heating of the vessel wall to cool the vessel wall, and applying acoustic energy to the liquid sample to disrupt a boundary layer of the liquid sample at the vessel wall so as to enhance heat transfer from the vessel wall to the liquid sample and to cool the liquid sample.

25. The method of claim 21, wherein the coupling medium includes liquid water and the acoustic energy source includes a transducer in contact with the liquid water.

26. The method of claim 21, wherein the acoustic energy is focused to form a focal zone of acoustic energy that is located at least in part at the vessel wall.

27. The method of claim 21, wherein the acoustic energy is focused to form a focal zone of acoustic energy that is located inside the vessel.

28. The method of claim 21, wherein the liquid sample increases in temperature at a rate of at least 50 degrees C. per ml per minute.

29. The method of claim 21, further comprising: subsequent to raising the temperature of the liquid sample above a temperature of the coupling medium, transferring heat from the vessel wall to the coupling medium via the heat exchanger so as to lower the temperature of the vessel wall; and subsequent to raising the temperature of the liquid sample, applying acoustic energy from the acoustic energy source to the liquid sample to disrupt the boundary layer of the liquid sample at the vessel wall to transfer heat from the liquid sample to the vessel wall and lower the temperature of the liquid sample.

30. A vessel for holding a liquid sample to be treated with acoustic energy, comprising: a vessel having a wall with an inner surface and an outer surface and defining an interior volume to hold a liquid sample; and a heat exchanger feature at the inner surface of the wall, the heat exchanger feature including physical structure that disrupts a boundary layer of the liquid sample at the vessel wall in response to sample movement caused by acoustic energy applied to the sample.

31. The vessel of claim 30, wherein the heat exchanger feature includes a plurality of raised areas on the vessel wall.

32. The vessel of claim 30, further comprising a heat exchanger feature on the outer surface of the wall.

33. The vessel of claim 30, wherein the interior volume is between 1 .mu.L and 100 milliliters.

---

Description

---

RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Application No. 61/368,410, filed Jul. 28, 2010, which is hereby incorporated by reference in its entirety.

BACKGROUND

[0002] 1. Field of Invention

[0003] Aspects described herein relate to acoustic treatment of samples, such as liquid material contained in a well of a microtiter plate or other similar vessel. In some cases, acoustic treatment of a sample may involve enhancing heat transfer between the vessel wall and the sample, such as through the disruption of a boundary layer at a vessel wall.

[0004] 2. Related Art

[0005] Analytical techniques for biological and chemical samples often require an extreme physicochemical preparatory step to enable the desired analysis to be fully achieved. For example, extraction/digestion of herbicides and pesticides from plant tissue may require organic solvents (e.g., alcohols) and elevated temperatures (e.g., 50 degrees C.). This requirement to elevate the temperature of a sample to aid extraction of a desired component or constituent of a sample is a commonly used technique. For example, many environmental sample analysis techniques require thermal energy to aid extraction. Another area whereby thermal energy is utilized to aid sample preparation is in microbial analysis; difficult cell wall disruption is aided by thermal energy.

[0006] Typically, transfer of thermal energy for such processes is achieved when heat is transferred from an area at higher temperature to a region of the sample at a lower temperature. For a biological or chemical sample contained in an isolated environment within a sample vessel, such heat transfer occurs by convection-based diffusion processes (Brownian motion and eddy diffusion) and advective fluid bulk transport (larger-scale current flow) processes. This is inherently a slow process and is exacerbated as the sample volume is increased (i.e., where the volume increases at a greater rate than the contact thermal surface area).

[0007] For example, a standard extraction/digestion process often used with sample slurries employs a combination of a stirring magnetic field to rotate a magnetic stir-bar in the sample fluid contained in a glass vessel and a hot plate to heat the vessel. The stir-bar imparts large scale currents, which ideally uniformly transfer the heat at the vessel wall to the entire fluid. An alternative means to transfer thermal energy is to use focused microwave techniques for biological and chemical processing. Indeed, even with closed vessel microwave heat exchange techniques, a magnetic stir-bar is utilized to impart large scale currents in the sample to be processed.

SUMMARY

[0008] In accordance with aspects of the invention, control of acoustic energy enables both heating of the vessel wall to heat a sample and disruption of a boundary layer of a sample liquid at the vessel wall to enhance heat transfer between the vessel wall and the sample. In other words, acoustic peak positive and peak negative zones may impart fluid movement for large-scale current formation as well as heating of the vessel wall. Heating of the vessel wall may be caused by an intrinsic acoustic impedance mismatch between materials (e.g., between the vessel wall and a surrounding acoustic coupling medium) such that a portion of the acoustic compression/rarefaction energy is absorbed by the vessel wall. The acoustic energy may also cause portions of the sample located at the vessel wall to flow, thereby enhancing heat transfer from the vessel wall to the sample. As a result, both mixing and heating of the sample can be performed without physically contacting the sample with any structure aside from the vessel. Also, some processes may benefit from exposing the sample to both elevated pressures and temperatures (i.e., pressures and temperatures above ambient). Aspects described herein may be useful with such processes since the sample may be both thermally heated as well as exposed to elevated pressures by way of cavitation or other conditions caused by the acoustic energy.

[0009] In one aspect of the invention, a method for acoustic treatment of a sample contained in a vessel includes providing a vessel containing a liquid sample where the vessel has a wall in contact with the liquid sample. The vessel wall may include a heat exchanger on an inner surface that is in contact with the liquid sample and/or a heat exchanger on an outer surface of the wall that is in contact with an acoustic coupling medium. The heat exchanger on the inner and/or outer surfaces may take a variety of forms, such as fins, bumps, grooves and/or other physical features that help increase a surface area of the vessel wall in contact with the sample or a coupling medium. The heat exchanger features at the inner surface of the vessel may also, or alternately, be arranged to help disrupt a boundary layer of the liquid sample at the vessel wall, e.g., to help induce large scale mixing or other flow of the sample to enhance heat transfer. Thus, the method may further include applying acoustic energy from an acoustic energy source to the liquid sample to cause movement of portions of the liquid sample near the vessel wall, and using a heat exchanger on the inner surface of the vessel wall to interact with moving portions of the liquid sample and disrupt a boundary layer of the liquid sample at the vessel wall, such that disruption of the boundary layer enhances heat transfer between the vessel wall and the liquid sample.

[0010] Heat transfer between the vessel wall and the sample may be used to heat or cool the sample. For example, simultaneous with disrupting the boundary layer of the sample at the vessel wall, acoustic energy may be applied from the acoustic energy source to the vessel wall to heat the vessel wall and increase the vessel wall's temperature above a temperature of the liquid sample. As will be understood, heating the vessel wall causes heat transfer from the vessel wall to the liquid sample to raise the temperature of the liquid sample. In some embodiments, the temperature of the sample may be raised above a temperature of a coupling medium in contact with an exterior of the vessel. The temperature of the sample may be detected, e.g., by an infrared detector, and the acoustic energy controlled so as to maintain the sample temperature constant, or to vary the temperature of the sample.

[0011] In other embodiments, the sample may be cooled. For example, a temperature of the vessel wall may be below a temperature of the liquid sample, and the boundary layer may be disrupted to cause heat transfer from the liquid sample to the vessel wall so as to lower a temperature of the liquid sample. The vessel wall may be cooled in any suitable way, such as by transferring heat from the vessel wall to a coupling medium in contact with the vessel wall. In one embodiment, the coupling medium may be liquid water, although other liquid, solid and semi-solid materials may be used to couple acoustic energy to the vessel.

[0012] When heating or cooling the sample by transfer of heat between the vessel wall and a coupling medium, a heat exchanger at the outer surface of the vessel wall may be employed. The heat exchanger may include physical features on the vessel wall, such as fins, ribs, grooves, a metal element or other relatively highly thermally conductive member, and so on. Disruption of a boundary layer of the liquid sample at the vessel wall as discussed above may also assist in enhancing heat transfer between the sample and the vessel wall.

[0013] In another aspect of the invention, a method for acoustic treatment of a sample contained in a vessel includes providing a vessel containing a liquid sample where the vessel has a wall with an inner surface in contact with the liquid sample. A coupling medium, which may be a single material such as liquid water, or two or more materials, may be provided in contact with an outer surface of the vessel such that the coupling medium may transmit acoustic energy to the vessel. Acoustic energy may be applied from an acoustic energy source through the coupling medium to the vessel wall to heat the vessel wall and increase the vessel wall's temperature above a temperature of the liquid sample. As discussed above, for some embodiments, the acoustic energy may take advantage of impedance mismatches between the vessel wall and the coupling medium and/or the sample to heat the vessel wall. Simultaneous with applying acoustic energy to heat the vessel wall, acoustic energy may be applied from the acoustic energy source to the liquid sample to disrupt a boundary layer of the liquid sample at the vessel wall so as to enhance heat transfer from the vessel wall to the liquid sample and to raise the temperature of the liquid sample above a temperature of the coupling medium. In one embodiment, heating of the liquid sample may be performed at a rate of at least about 25 degrees C. per ml per minute. This rapid heating capability is unknown in the prior art, and may be enabled by the use of a heat exchanger or other element to disrupt the boundary layer of the liquid sample at the vessel wall. That is, by physically disrupting the boundary layer, more effective sample flow or other movement may be caused, which results in more efficient heat transfer.

[0014] These and other aspects of the invention will be apparent from the following description and claims.

BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Aspects of the invention are shown and described with reference to illustrative embodiments and the following drawings, in which like numerals reference like elements, and wherein:

[0016] FIG. 1 shows a schematic diagram of an acoustic treatment system in accordance with an aspect of the invention;

[0017] FIG. 2 shows a schematic cross sectional diagram of a vessel having a heat exchanger element at an inner surface in a illustrative embodiment; and

[0018] FIG. 3 shows a vessel having a heat exchanger at an outer surface in an illustrative embodiment.

DETAILED DESCRIPTION

[0019] Although aspects of the invention are described with reference to embodiments in which acoustic energy is used to heat and/or cool a sample, the sample may be subjected to other treatments or other processes by the acoustic energy. For example, the acoustic energy may also be suitable, or be adjusted, to cause other effects in the liquid, such as fluidizing the sample, mixing the sample, stirring the sample, catalyzing the sample, disrupting the sample (such as shearing or fragmenting DNA molecules or other compounds, lysing cells, etc.), permeabilizing a component of the sample, enhancing a reaction in the sample (such as binding of material to the material supports), causing crystal growth in the sample (e.g., by nucleating crystal growth sites and/or enhancing the rate of crystal growth), preparing formulations (e.g., suspensions and/or emulsions suitable for therapeutic use), causing flow in a conduit, and/or sterilizing the sample. Thus, the acoustic energy may be used for other purposes than merely heating and/or cooling a sample. In other embodiments, the acoustic energy may facilitate chemical or other reactions in the liquid, which generate an end product that is to be separated from the liquid and other substances in the liquid, e.g., using beads or other structures that bind to the end product to be separated. In addition, under the applied acoustic energy, a controlled active turbulent regime may exist, whereby the collision frequency between binding partners in the sample and on beads or other structures is increased. This actively controlled turbulence may accelerate desired processes, as opposed to passive diffusion dominated processes of paramagnetic or other currently available bead products.

[0020] The samples can be treated in any suitable vessel provided that the vessel in at least some embodiments includes one or more aspects of the invention. Vessels can be sealed for the duration of the treatment to prevent contamination of the sample or of an environment outside of the vessel, and 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, optionally 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. 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. Thus, a sample vessel 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.

[0021] Any suitable sample material can be included in a vessel, and the sample may include any suitable combination of a liquid (such as a solvent), a solid material (such as pieces of bone, tissue or plant materials), a dissolved material (such as a salt) and so on. Some example materials that may be included in a sample are DNA, RNA, nucleic acids, or other genetic material, antibodies, receptors and/or ligands associated with cellular functions, proteins, polymers, amino acid monomers, an amino acid chain, enzymes, nucleic acid monomers or chains, saccharides or polysaccharides, lipids, organic or inorganic molecules, vectors, plasmids, pharmaceutical agents, compositions suitable for crystal growth, prions, bacteria, and/or viruses. This is not intended to be an exhaustive list, but rather to provide a few examples of sample material that may be used with aspects of the invention.

[0022] FIG. 1 shows a schematic diagram of an acoustic treatment system 100 that incorporates one or more aspects of the invention. In this illustrative embodiment, the system 100 includes an acoustic transducer 1 that is arranged to emit sonic energy through a coupling medium 2, such as a liquid (e.g., water, organic solution, etc.) held in a container 3 or a solid material (e.g., elastomeric material, gel, silicone, rubber, etc.) in contact with the transducer 1, and form a focal zone 11 of acoustic energy near or at a vessel 4. The acoustic energy at the focal zone may be suitable to cause heating, mixing, cavitation or other effects in a sample 6 located in the vessel 4. Cavitation or other conditions induced by acoustic energy at the focal zone may create localized relatively high pressure (and/or low pressure) conditions that may be useful in enhancing reactions in sample materials. The vessel 4 may have an interior volume of any suitable size, e.g., between 1 .mu.L and 100 milliliters.

[0023] A controller 5 may provide suitable control signals to the transducer 1 to generate desired acoustic energy, and control the relative position of the vessel 4 and the transducer 1 (e.g., in 3 dimensions) so that the sample 6 in the vessel 4 may be suitably positioned relative to the focal zone 11. Further details regarding an illustrative embodiment for an acoustic treatment system 100 are provided below, and in U.S. Pat. No. 6,948,843, which is incorporated herein by reference in its entirety. For example, the focal zone 11 may have a spherical, egg-like, or elongated rod-like shape, may include two or more focal zones or focal lines (e.g., focal zones with high aspect ratios), and so on.

[0024] In accordance with an aspect of the invention, the vessel 4 may include one or more heat exchanger features that are located in contact with the sample 6 and/or in contact with the coupling medium 2. When used at the inner surface of the vessel, the heat exchanger features can enable rapid heating of the sample, e.g., by enabling the disruption of a boundary layer of the sample at the vessel wall. Generally, the boundary layer may be considered herein as a layer of fluid immediately adjacent to a solid surface where certain effects (e.g., due to viscosity) arising from the presence of the solid surface play a non-negligible role. For example, a boundary layer may be a fluid layer adjacent a vessel wall that, in the absence of acoustic mixing/agitation, remains relatively stagnant, substantially does not transfer heat between the vessel wall and the fluid by convection, and instead transfers heat between the vessel wall and the fluid by radiation and/or conduction. When the boundary layer is sufficiently disrupted (e.g., by focused acoustic treatment), convective heat transfer between the vessel wall and the fluid occurs more freely. In some embodiments, for a vessel lacking a heat exchanger or similar feature at the inner wall, the boundary layer of sample at the vessel wall may remain undisturbed, essentially forming a region that behaves as a blanket of insulation that forces heat transfer by radiation or conduction processes only. In contrast, the heat exchanger features in accordance with an aspect of the invention at the inner wall of a vessel allow the acoustic energy to disrupt this boundary layer, thereby enabling convective heat transfer in addition to radiation and conduction modes.

[0025] Disruption of the boundary layer enabled by a heat exchanger feature creates large scale flow at the vessel wall and thus permits rapid heat transfer between the sample and the vessel. In cases where the vessel wall is at a higher temperature than the sample, the sample can be heated quickly, particularly where the vessel wall is being heated by acoustic energy. FIG. 2 shows a cross sectional view of a vessel 4 that includes heat exchanger features 7 in the form of an array of raised areas on the inner surface of the vessel wall. In this embodiment, the raised areas are arranged in a regular pattern of individual bumps that extends around the inner periphery of the vessel 4 and along at least part of the length of the vessel 4. These bumps 7 cause turbulence in flow occurring near the vessel wall, thereby breaking up a relatively stagnant boundary layer that might otherwise form. This breakup induces improved convective heat flow, allowing the sample to be heated or cooled more rapidly. The inventor has found that these features can enable extremely rapid heating of at least 25 degrees C. per milliliter of liquid per minute (degrees C. per ml per min). Heating this rapid is unknown in prior art applications that do not involve focused acoustics and one or more aspects of the invention.

[0026] The heat exchanger features 7 can be formed in any suitable way such as by molding, thermoforming, machining, etching, applying with an adhesive, and so on. For example, the heat exchanger 7 may be formed as part of a sleeve that is inserted into the vessel and bonded (e.g., with an adhesive, application of pressure, mechanical fit, etc.) to the inner wall. In another embodiment, the heat exchanger 7 may be molded integrally with the vessel wall. In addition, the shape, size and arrangement of heat exchanger features may be arranged in any suitable way. In the embodiment of FIG. 2, the heat exchanger features have a mesa-type shape, but may be arranged as fins, rods, smooth bumps, grooves, holes, pits, tabs, and others. Also, in this embodiment, the raised areas have a size of about 1 sq. millimeter, a height of about 100 micrometers and are separated from each other by a spacing of about 3 millimeters, but other sizings and spacings are possible. For example, the size, shape and/or space between features may be varied according to a frequency or set of frequencies used to treat the sample 6. In one embodiment, a variety of differently sized and spaced features may be used so that different sets of features may selectively interact with acoustic energy within a certain frequency range. That is, features of a first size/shape/spacing may interact most strongly with acoustic energy in a first frequency range, features of a second size/shape/spacing may interact most strongly with acoustic energy in a second frequency range, and so on. As a result, the different features may be activated at different times, e.g., if the sample 6 is treated with a sweep of varying frequency acoustic energy.

[0027] Heat exchanger features may be formed as positive features that extend from the vessel wall into the vessel and/or negative features that extend into the vessel wall. Different types of heat exchanger features may be used together, such as an array of bumps combined with an array of grooves. In short, the heat exchanger features may be arranged so as to maximize boundary layer disruption for one or more particular applications. Since different applications may involve different materials in the sample and/or different sample viscosities, the heat exchanger features may be arranged to work best with a specific sample viscosity range and/or particle sizes.

[0028] As noted above, a vessel 4 may include heat exchanger features at an inner surface of the vessel wall or at an outer surface of the wall. FIG. 3 shows another embodiment in which a vessel 4 includes heat exchanger features 8 on an exterior of the vessel. In this embodiment, the heat exchanger features 8 are arranged as longitudinal fins that extend along a length of the vessel. In contrast to the heat exchanger features 7 at the interior of the vessel, the heat exchanger features 8 on the exterior of the vessel need not necessarily function to disrupt a boundary layer of a coupling medium or other liquid at the exterior of the vessel. Instead, heat exchanger features 8 at the vessel exterior may function to help increase surface area and heat transfer to a liquid, solid or semi-solid coupling medium (such as water, a silica material, and/or a silicone rubber). By exchanging heat with the coupling medium, the vessel can be heated and/or cooled so long as there is a temperature difference between the vessel and the coupling medium. As discussed above, heat transfer between the vessel and the sample can heat and cool the sample, and thus the coupling medium can be used to cool and/or heat the sample in certain circumstances. By providing heat exchanger features 8 on the vessel exterior, heat transfer between the vessel and the coupling medium can be better controlled, allowing for more accurate and efficient thermal cycling treatments of the sample to be performed.

[0029] As with the heat exchanger features 7 at the vessel interior, the heat exchanger features 8 can be arranged in any suitable way, with any suitable size, shape and/or configuration. Although the FIG. 3 embodiment shows the heat exchanger features 8 in the form of longitudinal fins, the heat exchanger features may include bumps, grooves, pits, circumferential or spiral fins (e.g., having a washer-like shape), plates, mushroom-like structures, studs, and others. The heat exchanger features 8 may be formed unitarily with the vessel (e.g., molded into the vessel wall), attached to the vessel wall (e.g., by an adhesive, sonic welding, or other) and so on. For example, in one embodiment, heat exchanger features 8 may be formed on a sleeve (such as a highly conductive metal sheath) that is slid over the vessel and bonded in place. In other embodiments, the heat exchanger features may be attached to the vessel using an interference or friction fit, such as metallic washer-shaped elements that are pressed onto the vessel wall such that the hole of the washer element fits tightly to the vessel outer surface. The heat exchanger may have portions that extend through the vessel wall, such as metallic stud elements that extend from outside the vessel wall, through the wall and into the vessel interior. In one embodiment, such heat exchanger features may be molded with a plastic material to make the vessel. For example, the metallic studs may be mounted in a mold and molten plastic injected so that the studs are formed integrally with the vessel and extend from inside to outside of the vessel. In one embodiment, such studs or similar elements may form both heat exchanger features at the inner surface of the vessel wall and heat exchanger features at the outer surface of the vessel wall.

[0030] When using a vessel in accordance with aspects of the invention, the temperature of the external environment of the sample vessel (e.g., the coupling medium) may be below the temperature of the sample during a treatment process. This arrangement enables the sample to be intermittently elevated in temperature for a desired process. For example, a sample in a polypropylene plastic tube and cap may initially be at 4 degree C. with the tube placed in a 96 tube rack. A focused acoustic field may be directed to the sample, which is contained in one of the tubes in the rack. During an acoustic dose, the internal temperature of the sample may be increased to 50 degree C. within seconds (e.g., less than 10 seconds). If the sample is initially frozen, this thermal energy may be used to quickly thaw the frozen sample. In accordance with an aspect of the invention, only one of the samples in the rack may be thawed while other samples remain frozen. This would be of benefit if the rack of samples (e.g., 96 tubes) were at -20 degree C., but only one sample was required to be thawed for processing. Rapid heating enabled by aspects of the invention has been found by the inventor to be significantly faster than other prior processes. For example, compare a process of thawing a biological fluidic sample (e.g., serum) that is initially at (-70) degrees C. in which the sample is placed in a water bath at 20 degree C. to a process in accordance with aspects of the invention. Simply placing the sample in a 20 degree C. water bath typically requires several minutes before the sample reaches a temperature at 20 degree C. However, with an applied acoustic field and heat exchanger elements used with the vessel, a sample thaw may occur within 10 seconds even with the coupling medium at a relatively lower temperature of 5 degrees C.

[0031] In other embodiments, the temperature of the external environment of the sample vessel (e.g., the coupling medium temperature) may be elevated above the sample temperature, at least initially. In this situation, a rise in temperature of the sample, if desired, may be further accelerated. For example, a -70 degree C. frozen sample may be placed into a water bath of 20 degrees C. and an acoustic dose applied to the vessel. As the vessel wall is heated by the acoustic energy, the fluid motion turbulence generated by the acoustic energy and a heat exchanger in the vessel further aids the heat transfer from the vessel wall and the coupling medium to the sample. Similar is true where the sample is to be cooled where the sample temperature is higher than the coupling medium. Thus, the heat transfer process may be accelerated for both heating and cooling of the sample by appropriate setting of the coupling medium temperature. This may be of value in thermal cycling of biological processes, such as thermo-stabile enzymes.

[0032] The controller 5 may be arranged to control the transducer 1 in any suitable way, e.g., generate a variety of alternating voltage waveforms to drive the transducer 1. For instance, a high power "treatment" interval consisting of about 5 to 1,000 sine waves, for example, at 1.1 MHz, may be followed by a low power "convection mixing" interval consisting of about 1,000 to 1,000,000 sine waves, for example, at the same frequency. (Although there is a short time period separation between treatment and mixing intervals, the intervals are referred to herein as occurring simultaneously, i.e., acoustic energy to cause heating is said to be applied simultaneously with acoustic energy to cause mixing.) It is during the convective mixing interval that heat exchanger elements in the vessel may maximally assist in disrupting the boundary layer at the vessel wall. "Dead times" or quiescent intervals of about 100 microseconds to 100 milliseconds, for example, may be programmed to occur between the treatment and convection mixing intervals. Also, the focal zone 11 may be arranged in any suitable way, e.g., may be small relative to the dimensions of the vessel 4 to avoid heating of the treatment vessel during some intervals, or may be larger than the vessel 4. In one embodiment, the focal zone 11 may have a width of about 2 cm or less, a height of about 6 cm or less and a length of 5 cm or more. In another embodiment, the focal zone 11 may have an ellipsoidal shape, with a width or diameter of about 2 cm or less and a length of about 6 cm or less.

[0033] Acoustic energy in the focal zone 11 may generate a shock wave that is characterized by a rapid shock front with a positive peak pressure in the range of about 15 MPa, and a negative peak pressure in the range of about negative 5 MPa. This waveform may be of about a few microseconds duration, such as about 5 microseconds. If the negative peak is greater than about 1 MPa, cavitation bubbles may form in liquid in the sample. Cavitation bubble formation may also be dependent upon the surrounding medium 2, the vessel material, or other features. For example, glycerol is a cavitation inhibitive medium, whereas liquid water is a cavitation promotive medium. The collapse of cavitation bubbles may form "microjets" and turbulence that impinge on the surrounding material. These cavitation bubbles may contribute to sample liquid movement during a treatment. Moreover, the localized high and low pressure regions may expose portions of the sample to suitable pressures and temperatures that are useful for causing some chemical reactions or other results.

[0034] In the embodiments shown, the acoustic energy is transmitted from the transducer 1 to the vessel 4 through a medium 2, such as water. However, other media or combinations of media may be used, such as a solid or semi-solid material and others. For example, the transducer 1 may be mated to a solid silica-based material that conducts acoustic energy toward the sample vessel. A semi-solid material, such as a silicone rubber or gel, may be used to couple the silica material to the vessel. The water bath or other acoustic coupling media (e.g., silicone rubber) may be at room temperature and the sample may be contained in a vessel which readily transfers heat (e.g., borosilicate glass), but allows the acoustic energy to couple directly with the internal sample for heat transfer. For example, a 20% glycerol sample will be more sensitive to acoustic energy-mediated temperature elevation than a 2% glycerol sample. In this embodiment, the vessel wall may be more transparent to acoustic energy, and thereby resulting in the sample or sample constituents absorbing the acoustic energy and impart thermal energy transfer directly to the sample. An example of a vessel wall material with desired acoustic properties is the low density, transparent thermoplastic polymer of methylpentene monomer units (polymethylpentene or TPX).

[0035] The geometry and material choice of the vessel wall may also affect the performance of the non-contact, acoustic treatment. In addition, the internal vessel volume and the ratio of sample to headspace will also affect the performance of the device. For example, a 1.5 milliliter conical polypropylene tube with 1.0 milliliter of sample when placed into a 0.5 MHz focused acoustic field converging on the cone of the tube would enable the internal, starting temperature of 20 degree C. (external water bath temperature) to be elevated to 90 degree C. in less than 120 seconds at a high acoustic dose. The temperature may quickly be lowered to 20 degree C. with a lower acoustic dose to dissipate the thermal energy.

[0036] Many types of acoustic systems may be used to generate the appropriate wave-train to impart the thermal energy transfer. For example, an unfocused acoustic source (15 kHz) directed toward the vessel would result in the vessel wall temperature rise, which would thereby heat the internal fluidic sample. Alternatively, a focused acoustic source (e.g., 0.5 MHz) may also be used. In both situations, a feedback loop algorithm may be utilized to automate and control the process, e.g., monitoring the external temperature of the vessel wall may indirectly indicate the appropriate dose to be applied to the sample. In one embodiment, the apparatus may have an external non-contact infrared meter monitoring the external temperature of the sample vessel. For example, during an acoustic extraction dose, the vessel wall temperature will increase and the fluidic sample will be turbulent. The turbulence will effectively transfer the temperature throughout the sample and thereby enable external thermal measurements to provide an indication of internal temperature. This is particularly valid if the sample is thoroughly washing the internal walls of the vessel during the acoustic dose. Thus, a heat exchanger 7 at the vessel inner surface may enable more accurate temperature measurement of the sample.

[0037] Many biological and other materials can be treated according to aspects of the invention. For example, such materials for treatment include, without limitation, growing plant tissue such as root tips, meristem, and callus, bone, yeast and other microorganisms with tough cell walls; bacterial cells and/or cultures on agar plates or in growth media, stem or blood cells, hybridomas and other cells from immortalized cell lines, and embryos. Additionally, other biological materials, such as serum and protein preparations, can be treated with the processes of the invention, including sterilization.

[0038] Many binding reactions can be enhanced with treatments in accordance with aspects of the present disclosure. Binding reactions involve binding together two or more molecules, for example, two nucleic acid molecules, by hybridization or other non-covalent binding. Binding reactions are found, for example, in an assay to detect binding, such as a specific staining reaction, in a reaction such as the polymerase chain reaction where one nucleotide molecule is a primer and the other is a substrate molecule to be replicated, or in a binding interaction involving an antibody and the molecule it binds, such as an immunoassay. Reactions also can involve binding of a substrate and a ligand. For example, a substrate such as an antibody or receptor can be immobilized on a support surface, for use in purification or separation techniques of epitopes, ligands, and other molecules.

[0039] In certain embodiments, temperature, mixing, or both can be controlled with ultrasonic energy to enhance a chemical reaction. For example, the association rate between a ligand present in a sample to be treated and a binding partner on a bead or other support in the sample can be accelerated. In another example, an assay is performed where temperature is maintained and mixing is increased to improve association of two or more molecules compared to ambient conditions. It is possible to combine the various aspects of the process described herein by first subjecting a mixture to heat and mixing in order to separate a ligand or analyte in the mixture from endogenous binding partners in the mixture. The temperature, mixing, or both, are changed from the initial condition to enhance ligand complex formation with a binding partner relative to ligand/endogenous binding partner complex formation at ambient temperature and mixing. Generally, the second temperature and/or mixing conditions are intermediate between ambient conditions and the conditions used in the first separating step above. At the second temperature and mixing condition, the separated ligand may be reacted with the binding partner.

[0040] Focused sonic fields can be used to enhance separations. As noted elsewhere, sonic foci can be used to diminish or eliminate wall effects in fluid flow, which is an important element of many separation processes, such as chromatography including gas chromatography, size exclusion chromatography, ion exchange chromatography, and other known forms, including filed-flow fractionation. The ability to remotely modulate and/or reduce or eliminate the velocity and concentration gradients of a flowing stream is applicable in a wide variety of situations, such as those described in relation to FIG. 2.

[0041] Sonic energy fields can be used to enhance reaction rates in a viscous medium, by providing remote stirring on a micro scale with minimal heating and/or sample damage. Heat exchanger features in a vessel may be useful in promoting micro and larger scale stirring whether with or without significant heat transfer. Likewise, any bimolecular (second-order) reaction where the reactants are not mixed at a molecular scale, each homogenously dissolved in the same phase, potentially can be accelerated by sonic stirring. At scales larger than a few nanometers, convection or stirring can potentially minimize local concentration gradients and thereby increase the rate of reaction. This effect can be important when both reactants are macromolecules, such as an antibody and a large target for the antibody, such as a cell, since their diffusion rates are relatively slow and desorption rates may not be significant.

[0042] These advantages may be realized inexpensively on multiple samples in an array, such as a microtiter plate. The use of remote sonic mixing provides a substantially instantaneous start time to a reaction when the sample and analytical reagents are of different densities, because in small vessels, such as the wells of a 96 or 384 well plate, little mixing will occur when a normal-density sample (about 1 g/cc) is layered over a higher-density reagent mixture. Remote sonic mixing can start the reaction at a defined time and control its rate, when required. Stepping and dithering functions may allow multiple readings of the progress of the reaction to be made. The mode of detecting reaction conditions can be varied between samples if necessary. In fact, observations by multiple monitoring techniques, such as the use of differing optical techniques, can be used on the same sample at each pass through one or more detection regions.

[0043] By focusing and positioning sonic energy near a wall of a vessel, e.g., at heat exchanger features, many local differences in the distribution of materials within a sample and/or spatially-derived reaction barriers, particularly in reactive and flowing systems, can be reduced to the minimum delays required for microscopic diffusion. Put differently, enhanced mixing can be obtained in situations where imperfect mixing is common. For example, if sonic energy is focused in, on, or near the wall of the vessel, near the fluid/wall boundary, then local turbulence can be obtained without a high rate of bulk fluid flow. Excitation of the near-wall fluid in a continuous, scanned, or burst mode can lead to rapid homogenization of the fluid composition just downstream of the excited zone, e.g., a short distance away from a boundary layer at a heat exchanger feature.

[0044] This effect is useful in several areas, including chromatography; fluid flow in analytical devices, such as clinical chemistry analyzers; and conversion of the fluid in a pipeline from one grade or type to another. Since most of the effect occurs in a narrow zone, only a narrow zone of the conduit typically needs to be sonically excited, and only the narrow zone need include heat exchanger features at the vessel wall. For example, in some applications, the focal zone of the sonic energy can be the region closest to a valve or other device which initiates the switch of composition. In any of these applications, feedback control can be based on local temperature rise in the fluid at a point near to or downstream of the excitation region.

[0045] Focused acoustics and heat exchanger features in accordance with aspects of the present disclosure may be useful for preparing formulations having a narrow particle size distribution. Such formulations may include suspensions and/or emulsions having particles that are submicron in size and may have applications for therapeutic use, such as delivery systems for bioactive agents (e.g., liposomes, micelles, etc.). Controlling heat transfer in a focused acoustic processing system using heat exchanger features contemplated may enhance the ability to suitably prepare formulations in an advantageous manner (e.g., repeatable, short processing times, high yield, etc.).

[0046] In some embodiments, enhancing heat transfer between the wall of a processing vessel and a fluid upon focused acoustic treatment may also be useful for initiating (e.g., forming nucleation sites) and augmenting (nano)crystalline growth. For example, crystalline particles may be formed as a suspension of particles (e.g., submicron in size) in a liquid solution. In some cases, though not required, (nano)crystalline particles prepared in accordance with aspects described herein may be useful for therapeutic delivery of bioactive agents.

[0047] While there has been described herein what are considered to be exemplary and preferred embodiments of the invention, other modifications and alternatives of the inventions will be apparent to those skilled in the art from the teachings herein. All such modifications and alternatives are considered to be within the scope of the invention.

* * * * *