Controller for an acoustic standing wave generation device in order to prevent clogging of a filter

Abstract

The invention relates to a method and apparatus to continuously monitor and control acoustic energy and vacuum pressure to maintain a net unidirectional flow of multi-phase heterogeneous fluids through a porous filter or membrane. The heterogeneous fluid may come form a variety of sources including biological sources such as, blood, bone marrow aspirate (BMA), adipose tissue (lipoaspirate), urine, saliva, etc.

Claims

1. An apparatus for separating a fraction from a fluid sample, the apparatus comprising: a filtration unit including at least one filter which divides the filtration unit into a prefiltration chamber for receiving the fluid sample and a post-filtration chamber for receiving a filtrate of the fluid sample; the filter movable from a horizontal orientation to a tilted orientation; a fluid barrier abutting a porous top surface of the filter; the filter configured to form a pool area on a lowered portion of the porous surface against the fluid barrier when the filter is in the tilted orientation; and a filter tilting device coupled to the filter and configured to tilt the filter from the horizontal orientation to the tilted orientation.

2. The apparatus of claim 1, further comprising: an electronic measuring device to measure a degree of filtration and for generating a signal when a predetermined degree of filtration has been achieved; wherein the filter tilting device is operable to tilt the filter in response to the signal from the electronic measuring device.

3. The apparatus of claim 2, wherein the electronic measuring device comprises at least one sensor configured for measuring an amount of fluid in the prefiltration chamber and/or in the postfiltration chamber.

4. The apparatus of claim 3 wherein the at least one sensor is configured for measuring a volume or level of fluid in the prefiltration chamber or in the post filtration chamber.

5. The apparatus of claim 3 wherein the at least one sensor is configured for measuring a weight or mass of fluid in the prefiltration chamber or in the post filtration chamber.

6. The apparatus of claim 3 wherein the at least one sensor is configured for measuring time elapsed while the apparatus is separating the fraction from the fluid sample.

7. The apparatus of claim 3 wherein the at least one sensor is configured for measuring a degree of turbidity of fluid in the prefiltration chamber.

8. The apparatus of claim 1, wherein the filter tilting device is manually operable by a user of the apparatus when it is determined that filtration has proceeded to a required degree.

9. The apparatus of claim 1, wherein the filter tilting device, once operated, does not allow the filter to be returned to or to be retained in the first orientation relative to the horizontal plane.

10. The apparatus of claim 1, wherein porous surface is configured to collect a film of a filtrand when the filter is in the horizontal orientation, and to allow the filtrand to flow on the porous surface into the pool area when the filter is in the tilted orientation.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) For a better understanding of the present invention and to show how it may be carried into effect, reference shall now be made by way of example to the accompanying drawings, in which:

(2) FIG. 1: A schematic showing the general operating principle of a filtration unit of the type incorporated in embodiments of the invention.

(3) FIG. 2: A photograph of an embodiment of the filtration unit according to the invention.

(4) FIG. 3: A photograph of an embodiment of the separation apparatus according to the invention.

(5) FIG. 4: A schematic of an embodiment of the separation apparatus according to the invention.

(6) FIG. 5: A photograph of the LCD user interface on the control unit.

(7) FIG. 6: A flow diagram showing a general operating principle of the controller and its connected components.

(8) FIG. 7: A chart showing the role of the controller PCB in controlling, monitoring and regulating vacuum pressure, fluid volume/load and acoustic energy.

(9) FIG. 8: A plot showing mass-frequency correlation in an embodiment of the present invention.

(10) FIG. 9: A plot showing the measured and calculated correlations for acoustic frequency against mass in an embodiment of the present invention for both human and porcine bone marrow aspirate (BMA).

(11) FIG. 10: An exploded view of a vibrating substrate suitable for use with embodiments of the present invention.

(12) FIG. 11: A schematic view of embodiment of the apparatus incorporating an automatic tilt mechanism in (a) a first configuration and (b) a second configuration.

(13) FIG. 12: A schematic diagram showing a relationship between tilt angle, hinge-to-spring distance and spring extension.

(14) FIG. 13: A flow diagram showing a decision-making process used in embodiments illustrated in FIGS. 11 and 12.

(15) FIG. 14: A photograph of an embodiment of the apparatus utilising an automatic tilt mechanism. Shown in the pre-load configuration.

(16) FIG. 15: A photograph of an embodiment of the apparatus utilising an automatic tilt mechanism. Shown in the tilted configuration.

(17) FIG. 16: A schematic view of an alternative embodiment of the apparatus incorporating an automatic tilt mechanism in a first and second configuration.

(18) FIG. 17: A schematic view of an embodiment of the apparatus incorporating a mechanical tilt mechanism.

(19) FIG. 18: A photograph of the embodiment illustrated in FIG. 17.

(20) FIG. 19: A schematic view of an alternative embodiment of the apparatus incorporating a mechanical tilt mechanism.

(21) FIG. 20: A schematic view of an embodiment of the apparatus incorporating a mechanical tilt mechanism.

(22) FIG. 21: A schematic view of an alternative embodiment of the apparatus incorporating a mechanical tilt mechanism.

(23) FIG. 22: A schematic view of an alternative embodiment of the apparatus incorporating a mechanical tilt mechanism.

(24) FIG. 23: A schematic view of an alternative embodiment of the apparatus incorporating a mechanical tilt mechanism.

(25) FIG. 24: A schematic view of an alternative embodiment of the apparatus incorporating a mechanical tilt mechanism.

(26) FIG. 25: Cell viability comparison between unprocessed and unprocessed human BMA using the apparatus of the invention.

(27) FIG. 26: Retention of TNC and equivalent fold-concentration as a consequence of filtration in the apparatus of the invention.

(28) FIG. 27: CFU-f and CFU-Ob comparison between processed and unprocessed BMA for a 9-fold volume reduction. From 25 human BMA samples mean CFU-f was 880/cc for unprocessed samples and 4190/cc for processed samples. Of these, 82% were CFU-Ob and 89% CFU-Ob for unprocessed and processed respectively.

(29) FIG. 28: Clinical efficacy of concentrated BMA from apparatus of the invention based on a non-unionfracture study by Hernigou

DETAILED DESCRIPTION

(30) FIG. 1: A schematic showing the general operating principle of a separator apparatus of a type incorporated in embodiments of the present invention in which the following reference numerals refer: 1. Filtration unit 2. Porous filter 3. Upper (pre-filtration) chamber for receiving fluid sample. 4. Fluid sample 5. Lower (post-filtration) chamber for receiving back-flushing fluid. 6. Fluid provided in the post-filtration chamber 7. Resonating substrate 8. Acoustic energy generating element 9. Vacuum draw (optional)

(31) The porous filter 2 separates a filtration unit 1 into two chambers; an upper (pre-filtration) chamber 3 into which a fluid sample 4 requiring cell separation is introduced and a lower (post-filtration) chamber 5 into which a fluid 6 capable of transmitting an acoustic standing wave is introduced. An acoustic element 8 is coupled to a substrate 7 which is located within and at the bottom of the lower chamber and which resonates in response to the acoustic generating element and generates a standing wave through the two fluid phases and the filter to agitate the sample. Simultaneously, a cyclic process of vacuum draw 9 causes movement of the sample downwards through the filter. Vacuum pressure, fluid flow rate and frequency of vibration are controlled by a controller (associated with appropriate pumps and valves. A concentrated fraction of desired larger cells is retained on top of the filter whilst smaller cells pass through the filter to a waste receptacle (not shown).

(32) In a specific embodiment of the invention the acoustic element is a speaker having a power of 0.4 W, resistance of 40, amplitude in the range of between about 4.2V to 7.36V peak to peak and a frequency range in the range of between about 300-700 Hz.

(33) FIG. 2: A photograph of illustrating the component assembly of an embodiment of the filtration unit of the invention in which the following reference numerals refer: 10. upper chamber 11. middle chamber 12. lower chamber 13. clamps to secure upper chamber and middle chambers 14. membrane filter 15. O-rings sealing to filter when the upper and middle chambers are clamped together 16. upper tissue sample reservoir within middle chamber 17. input into saline reservoir below filter 18. acoustic energy generating element 19. O-rings sealing to acoustic element 20. exit for acoustic element electrical connection

(34) FIG. 3: A photograph of a separation apparatus of a type incorporated in embodiments of the present invention in which the fluids in the pre- and post-filtration chambers are sequentially moved across the filter, and in which the following reference numerals refer: 21. Filtration unit (process chamber) 22. Control unit 23. LCD: Acoustic frequency 24. LCD: Vacuum pressure 25. Drip counter 26. Drip sensor cable 27. Pressure sensor 28. Signal volume 29. Acoustic frequency 30. Vacuum knob 31. Pressure sensor cable 32. Pump switch 33. Audio cable 34. Saline line (from syringe to process chamber) 35. Waste line (from process chamber to waste chamber) 36. Waste chamber

(35) This figure illustrates an apparatus which comprises a filtration unit 21 and a control unit 22. The control unit 19 can be programmed to control the vacuum pump (Koge KPV14A-6A) (not shown). An amplifier and signal generator chip built into the control unit allows the frequency and amplitude of the acoustic element (not shown) to be set via the PLC. The PLC also operates together with a load cell (not shown) so as to vary the applied acoustic energy as the volume of fluid above the filter (not shown) changes, in accordance with aspects of the present invention.

(36) FIG. 4: Schematic representation of a further embodiment of the apparatus of the invention and in which the following reference numerals refer: 37. Filtration unit 38. Acoustic energy generating element 39. Load cell 40. Acoustic sensor 41. Interactive LDC panelLCD user interface 42. Micro processor 43. Printed circuit board, PCB 44. Vacuum pump 45. Pressure sensor 46. Waste chamber

(37) The PCB 43 is programmed to switch the acoustic energy generating element 38 and vacuum pump 44 (Koge KPV14A-6A) on and off. It is also integrated with a pressure sensor 45 and an acoustic sensor 40 (e.g. microphone) to constantly monitor and adjust the working vacuum pressure and the acoustic energy to an optimum. The LCD interface 41 guides the user through the entire process/procedure with interactive flashing icons indicating what the user should do in each step. The entire system is powered up by a Power Source e.g. batteries.

(38) FIG. 5: Photograph of the LCD user interface on the control unit of the invention and in which the following reference numerals refer:

(39) Icons 47: input saline 48: input biological fluid 49: input required final volume 50: Processing 51. Required volume reached (processing completed) 52: Press set/next button 53: Fluid volume 54: Battery power indicator

(40) Action Buttons 55: Set/next button 56: Up and down button for adjusting fluid volume.

(41) In normal operation the separation chamber of the apparatus is initially free of fluid. The LCD interface will display Input Saline 47 and Input Biological Fluid Mixture 48 icons to indicate the user to deliver the fluids into apparatus. The volume of the biological fluid mixture added is registered by the load cell and displayed on the LCD 53. This will be followed by the Input Required End Volume 49 icon which can be set by using the Up and Down Buttons 56 on the panel. Once the required final volume is set the biological fluid mixture will undergo processing, which will be indicated by the Processing in Progress 50 icon. During processing, the acoustic element and the vacuum pump are switched on. The acoustic energy and the vacuum pressure applied will be constantly monitored and automatically adjusted as the processing fluid volume decreases. The acoustic energy has amplitude fixed at 11V and an amplifier signal voltage of less than 5V. The signal volume range from 2 to 6 and the frequency range from 350 to 650 Hz, this drives a standing wave through the fluid and the fluid observed to be in constant agitation. The negative vacuum pressure applied range from 0.2 to 0.3 psi to keep a net unidirectional flow of biological fluid through the filter into the waste chamber. Once the desired/entered end volume is reached, Process completed icon appears 51, and the PCB is permanently disabled with a kill command form the micro-processor. The processed biological fluid above the filter is then removed and is ready for use.

(42) The flow diagram of FIG. 6 shows a currently preferred operating principle for the control system of embodiments of the present invention, with FIG. 7 showing the role of the PCB in controlling, monitoring and regulating the vacuum pressure, fluid volume/load and acoustic energy.

(43) The separation apparatus of FIG. 4 has a load cell that measures the mass of the fluid and a microprocessor that controls the frequency of an acoustic actuator. The fluid mass above the porous filter in the separation chamber was recorded every 20 seconds, as well as the corresponding acoustic frequency at that time point.

(44) A representative mass-frequency profile is shown in FIG. 8 for the separation apparatus using porcine bone marrow. The measured data is best represented by the correlation:
y=733.12x(e0.1516) with an R.sup.2=0.9759.

(45) In practice, the generalised correlation would be applied within the microprocessor software, such that for a given measured fluid mass the appropriate frequency would be applied to the acoustic actuator in the separation apparatus.

(46) Another representative mass-frequency profile is shown in FIG. 9 for the separation apparatus of FIG. 8 using both human and porcine bone marrow aspirate (BMA). The measured data is best represented by the linear regressions:
y=3.052x+502.83 (human BMA)
y=3.0122x+533.35 (porcine BMA)

(47) As fluid processing progressed, the mass of fluid contained above the filter was registered on an LCD coupled to a load cell. Simultaneously, the frequency of the acoustic element was registered on an independent LCD display. These data were generated using the same device.

(48) The regressions show that irrespective of tissue type the same linear change in frequency correlates to the change in fluid volume. The data also suggests that for human tissue there is constant reduced offset in frequency of approximately 30 Hz.

(49) Various materials may be used as a loudspeaker cone/diaphragm, but the most common are paper, plastic and metal. The ideal material would be light (to minimise starting force requirements), stiff (to prevent uncontrolled cone motions) and well damped (to reduce vibrations continuing after the signal has stopped). In practice, the three criteria cannot be met simultaneously using existing materials. As a result, many loudspeaker diaphragms are made of some sort of composite material.

(50) FIG. 10 shows an exploded view of a substrate or soundboard 57 made of composite material that, when used as loudspeaker cone/diaphragm in combination with an acoustic energy generating element, is capable of delivering appropriate acoustic energy into the biological fluid. It is a composite panel with layered/bonded sandwich construction, consisting of a polycarbonate disc core 58 and two outer stainless steel skins 59 of specific thickness. The outer skins 59 are extremely strong and the core 58 is lightweight and very much weaker, but with the use of a suitable adhesive the benefits are realised. Details are shown in Table 1. This combination of materials gives the soundboard 57 a unique material stiffness and performance characteristic such that, when used as speaker cone/diaphragm in combination with an acoustic actuator, it generates fluid resonance through efficient acoustic energy delivery which in turn provides efficient filtering in the cell separation apparatus of embodiments of the present invention.

(51) TABLE-US-00001 TABLE 1 Detail Supplier Core 0.5 mm thick polycarbonate sheet Fibrefusion Ltd (Cornwall, UK) Skin 0.05 mm thick 304 stainless steel sheet Rayhome Ltd (Bury, UK) Adhesive Huntsman Araldite 2022 methyl Huntsman methacrylate (or Araldite 2015 epoxy) Advanced Materials Americas Inc. (Texas, USA) Manufacturing Heated platen lamination machinery Fibrefusion Ltd process (Cornwall, UK) Size 60 mm

(52) A current working embodiment of an alternative embodiment of the invention is schematically represented in FIG. 11 comprising hinged separation chamber 60 together and a PCB/microprocessor 61.

(53) The hinged separating chamber is a pop-up sub-assembly held in a preloaded position as described below:

(54) The hinged supporting platform 62 is a moulding that incorporates the separation chamber as well as keeping the separation chamber and the porous filter 63 in the horizontal position. It is designed to pop-up to desired tilt angle once the biological fluid processing is complete, thus allowing for maximum recovery of the processed fluid. The actuator spring 64 is located at the opposite end to the hinge 65 sandwiching between the hinged supporting platform 62 and the base 66. It provides a uniform elevation force on the hinged separating chamber. The spring is under compression when the assembly is in the preloaded position.

(55) A fusible filament 67 (e.g. polymer filament loop) is tethered at one end to the hinged separation chamber (opposite to the hinge), drawn taut and tethered to the filament retainers 68 at the other end. This action anchors the separation chamber with the spring compressed such that the pop-up sub-assembly is grounded and preloaded. The filament is in direct contact with the fusible resistor 69 which, when activated, melts the filament and thereby allowing the preloaded subassembly to pop-up once processing is completed. The filament retainers hold the filament within the assembly by providing a method of attaching the filament to the pivoting bodies, whilst maintaining the tension in the filament in the preloaded position. The base provides the grounding points and guides for the filament to run through:

(56) When the specified final volume is reached (i.e. processing completed) and recognised by the load cell of the separation chamber, it triggers the PCB/microprocessor to activate the fusible resistor such that the filament is melted and broken at the point of contact. Once the thread is broken, the compression springs serve to release the anchored separation chamber that then mechanically locks out into the desired tilt angle. This is shown in FIG. 11b.

(57) The pre-set tilt angle is determined by (1) the uncompressed actuator spring length and (2) the position of the spring relative to the hinge (pivot point). This is demonstrated in FIG. 12, showing the relationship:
Tilt angle =tan.sup.1(O/A)
where: =Tilt angle O=length of relaxed springlength of compressed spring A=distance between hinge and spring

(58) FIG. 13 is a flow chart showing a decision-making process used in embodiments of the present invention illustrated in FIGS. 11 and 12.

(59) FIGS. 14 and 15 illustrate an embodiment of the invention in its pre-load and automatically tilted configurations, respectively. The Figures show the base 70 which includes a display 71 and user controls 72, the hinged supporting platform 73, the separation chamber 74 an outlet port 75 to which a syringe (not shown) may be attached in order to take a sample of filtrand or residue, and input ports 76, 77. The hinged supporting platform and the separation chamber are preferably configured as a disposable unit incorporating the tether (not shown). Once the tether has been broken and the hinged platform has popped up into the tilted configuration of FIG. 15, the hinged platform cannot be locked back in the preload configuration of FIG. 14, thereby preventing accidental re-use of the unit, which might otherwise result in cross-contamination between clinical samples and/or patient tissue.

(60) FIG. 16 shows an alternative embodiment in which the automatic tilt mechanism has been redesigned by using the filament or tether 78 to release a simple trigger mechanism instead of holding the full force of the sprung pivoting section. With this arrangement, the tether 78 would be only under a small amount of tensionjust enough to overcome a small spring force. For example, a small pivoting trigger 79 (e.g. made from polypropylene with a living hinge) would be under tension from a small spring 80, and held in its set position by the tether. When in the preload position, the pivoting section (e.g. a hook depending from the hinged supporting platform), would snap into place. When the tether is released, the trigger would be released and the pivoting section would pop-up.

(61) FIGS. 17 to 24 show alternative embodiments of the apparatus utilising manually-operated tilt means.

(62) In FIG. 17, a tilt lever 81 is hingedly mounted to the base 82 of a separation device. The tilt lever 81 comprises a span portion 82 and a pair of arms 83 with hinge pins 84. The hinge pins of the arms are adapted for snap fitting into complementary hinge recesses (not shown) in the base. As illustrated, when fitted to the base, the tilt lever 81 can be moved by hand from a first position (step 1) in which it is substantially recessed in the base, to a second position (step 4) in which the span portion projects from a bottom of the base causing the device to assume a tilted orientation on a surface on which it is disposed.

(63) A pair of recesses (not shown) are provided in opposed side walls of the base to enable the tilt lever 81 to be accessed easily by a user's fingers. The resulting tilt angle is determined by the width and angle of the span portion 82 when in the second position.