WO2019050840A1

WO2019050840A1 - Specimen preparation device

Info

Publication number: WO2019050840A1
Application number: PCT/US2018/049354
Authority: WO
Inventors: George Mismas; Gregory KARABINOS
Original assignee: George Mismas; Karabinos Gregory
Priority date: 2017-09-05
Filing date: 2018-09-04
Publication date: 2019-03-14
Also published as: EP3679350A4; EP3679350A1

Abstract

A specimen preparation device comprising a device frame, a test tube support plate for securing a plurality of test tubes including a plurality of elongated magnets and a plurality of elongated stop fins, each fin positioned to support one of the rows of test tubes by the test tube base, the fins extending parallel with the elongated magnets, a specimen air flow duct for providing a passage of air around or through the specimen, a return air duct including a heating element positioned inside the return air duct, and a forced air flow duct having at least one fan positioned inside forced air flow duct. The at least one fan positioned adjacent a plurality of fan baffles for circulating air through and around the specimen air flow duct and the return flow air duct.

Description

SPECIMEN PREPARATION DEVICE

Background of the Invention

Typically in the analysis of body fluid or body tissue, a sample must be taken and prepared in order to analyze the sample. The present invention is used in the preparation of the sample so the sample may be analyzed by moving the prepared sample to a device used in the analysis.

In one example, the device may be used in the analysis of cancer cells. A biopsy may be performed to remove tissue from a portion of an organ where cancer may be suspected. The present invention may prepare the tissue to perform an analysis for the detection of cancer.

Summary of the Invention

Bearing in mind the problems and deficiencies of the prior art, it is therefore an object of the present invention to provide a device which prepares a plurality of tissue or body fluid samples for analysis with minimal user intervention. It is another object of the present invention to provide a device which allows preparation of a large sample of DNA, RNA, body fluid or tissue.

It is yet another object of the present invention to provide a device which uniformly heats DNA/RNA samples for replication.

It is still another object of the present invention to provide a device which provides uniform magnetic fields to separate a plurality of samples for cleaning.

It is another object of the present invention to use vortexing in the preparation of DNA, RNA, body fluid or tissue samples.

Still other objects and advantages of the invention will in part be obvious and will in part be apparent from the specification. Description of the Preferred Embodiment(s)

In the present application, the term "vortexing" is defined as rotating or gyrating something in a circular or whirling direction. The term may refer to a strictly circular direction or a circular direction which includes a horizontal component. The specimen preparation device according to the present invention may be used in the preparation for analysis of DNA/RNA samples, body tissue samples or body fluid samples. The device may additionally be used in the preparation of other samples.

Photo 1 shows a front view of the specimen preparation device with a front lower panel removed. The air inlet duct is shown in an open position to allow air to be drawn into the front duct. A tray for test tubes is shown in the upper portion of the device. Photo 2 shows a front right side perspective view with a reagent distribution system having the ability to distribute reagents and to rinse tissue or body fluid samples using pipettes shown above the table portion of the device.

Photos 3 and 4 show the test tube tray which may be vortexed or shaken relative to the table. Photo 5 shows the test tube tray with the table of the device removed in order to show how the test tubes are supported by parallel elongated stop fins. Photo 6 shows a rear view of the device with the electronic controls exposed.

Photos 7 and 8 show the vortexing motor and shaker belts. A motor shaft has disposed on an upper portion a sprocket or pulley which rotates a second sprocket or pulley with a motor belt. The second sprocket rotates a shaft near one cornet of the test tube tray, the shaft which includes a pair of shaker sprockets each attached to sprockets on adjacent corners of the test tube tray. The fourth corner may be driven by any one of the other sprockets. Photo 1 3 shows counter balance weights on two of the vortexing shafts for preventing vibration in the stationary portions of the specimen preparation device.

Photos 9 and 10 show the motor and the sensing device for opening and closing the inlet deflector. The outlet deflector may include similar components for operating the outlet deflector. Photo 1 1 shows the front deflector in a closed position while Fig. 12 shows the front deflector in the open position. The specimen preparation device includes a device frame which is only shown in the photos in order to show the components within the frame in the accompanying drawings. The robotic substance transfer mechanism is shown in photos 1 and 2 and is disposed on the upper table of the device. The robotic substance transfer mechanism contains a fluid delivery probe and a fluid delivery conduit assembly or disposable tip which is in turn mounted on a support assembly which provides motion in 3

dimensional space. The fluid delivery probe is moved by motors disposed within the transfer mechanism housing. A fluid reservoir is connected by tubing to one port of a 3- way fluid valve. A second port of the 3-way valve is connected to a mechanism which can extract a measured amount of fluid from the fluid reservoir and dispense the fluid to another portion of inert tubing or dispensing tip. The pipette is mounted above a test tube. A third port of the 3-way fluid valve is attached to a flow control device, controlling the flow of the fluids from the reservoir to the test tubes. The 3-way valve may be directly connected to a syringe or collapsible bellows and in turn to a piston. The piston maybe controlled by rotation of a motor.

The lower portion of the specimen preparation device and the test tube portion are shown in a top left front perspective view of Fig. 1 A (forced air flow duct and return ducts removed), a top right rear perspective view of Fig. 1 B (the forced air flow duct removed), and a bottom left front perspective view in Fig. 2 (forced air flow duct removed), includes a test tube support plate for securing a plurality of test tubes in a vertical position, the test tubes being in an array of parallel rows. Fig. 2 also shows one of the four circulation fans attached to the fan bracket directly below the fan baffles, the other 3 fans removed for clarity and the four fans removed from all other figures for clarity. Fig. 3 shows a rear right top sectional view of the specimen preparation device and Fig. 4A is a left front bottom perspective view with the bottom plate and rear duct removed. Figs. 4B - 4D have various portions of the duct removed and more specifically Fig. 4 B shows the front duct having a screen for filtration or for providing more consistent air movement while Fig. 4C has the screen removed to show a clear view of the fan baffles. The figures show the specimen preparation device with an outer shell removed, the specimen preparation device including test tubes for containment, preparation and analysis of the specimens. The test tubes are disposed in a test tube rack in parallel rows. The test tubes include a cylindrical wall and a semispherical base at the bottom end of the cylindrical wall. As shown in Fig. 4D with the forced air and return air ducts having their respective side covers included, the air flowing through the specimen preparation device when the inlet and outlet are in the closed position forms a closed system where the air in the system is not exchanged with fresh or ambient air. The shaker bearing blocks may additionally have sides that may guide the air flow at the bottom portion of the specimen preparation device when the controllable air deflectors are in the closed position. There may be areas in the air flow path that are not tightly sealed to the ambient air, but the air flow produced by the fan does not pressurize the air to the degree that tight seals are necessary and a small amount of air leakage is not

problematic. There may be some seals in the system as shown in Fig. 1 9, but they are primarily used to seal large gaps between components.

Figs. 5-7 show the shaker assembly for vortexing the test tubes, a portion of which are the vortexing motor and shaker belt shown in Photos 7 and 8. The shaker assembly allows the contents of the test tubes to be mixed using a circular motion. The shaker assembly is attached to a stationary shaker bearing block which is mounted to the frame of the specimen preparation device. The shaker shafts rotate at one end within the shaker bearing block and at the opposite end within an upper and lower bearing retainer on the shaker plate. The upper portion of the shaker shaft is an offset shaft portion which moves the shaker plate in a circular direction as the shaft rotates. A motor connected to at least one of the shafts rotates the shaft and the sprockets on each of the shaker shafts rotate the remaining shafts by way of shaker belts connecting each of the shaker sprockets. There is a shaker assembly at each corner of the shaker plate to produce a smooth circular pattern of movement to the shaker plate.

The shaker plate shown in Figs. 1 A and 1 B is attached to the elevator guide rod and the elevator guide rods are mounted in the magnet elevator bearings such that the elevator guide rods may vortex within the bearing, allowing the magnet elevator guide rods to move in a vertical direction while moving in a circular direction. The shaker plate, elevator guide rods, test tube base air baker finish cover and rack top with test tubes are therefore able to vortex along with the magnets, magnetic array support bracket and test tube stops relative to the stationary frame and table top.

A volume of air may be circulated along the fins, heating the test tubes to a specified temperature, maintaining the specified temperature, cooling the volume of air and then repeating the heating and cooling process for a specified number of cycles. A screen may be provided to aid in providing uniform heating or in the filtration of particles in the circulated air.

The specimen preparation device includes an inlet and outlet deflector movable to a first position wherein heated air is circulated through the device without the

introduction of additional air to a second position wherein air is drawn in through the inlet duct, moved across the test tubes and is forced out through the outlet duct, allowing the test tubes to cool.

The tissue samples within the test tubes may be combined with iron filings or iron powder, the tissue sample being attracted to the iron filings during the shaking process whereby the iron filings are attracted to the sides of the test tube when the magnets are moved to the second positon and aspiration of the fluid and tissue in the test tube may occur by removing the fluid, adding a cleaning fluid to the test tube and with or without shaking the tube, removing the cleaning solution to leave substantially only the tissue sample attached to the iron filings. The specimen preparation device includes a system for heating and cooling the specimen samples, the system shown in Figs. 8 and 9 of the drawings. The air flow through the device is shown by the air flow arrows 70. The specimen preparation device shown has the outer panels removed to show the internal components and how the air flows through and around these components. Fig. 9 has the shaker bearing blocks and one side of the deflector end plates additionally removed for clarity.

Figs. 10, 1 1 A and 1 1 B show a detailed drawing of the positioning of the magnets on the magnet array mounting bracket (FIG. 1 1 ) in relation to the test tube support plate and the elongated slots in the test tube support plate. The test tube support plate is in a second positon wherein the magnets are adjacent the test tube cylindrical sides. In the drawing, the test tube rack plate, air baker cover magnet holder bottom, and all but three test tube rows and stops are removed to show the mounting portion of the magnet array mounting bracket. Additionally, only one magnet is shown for clarity. In a first position, the elongated magnets are horizontally between the rows of test tubes and vertically below level of the test tube bottoms. In the second position, the bottoms of the magnets are substantially level with the top surface of the test tube support plate and the magnets are adjacent the test tube cylindrical sides. In this configuration, there is a magnet on each side of the test tube, forcing the iron filings and corresponding tissue sample to be forged against the sides of the test tube and allowing for the cleansing solution to be added and removed without removing the iron filings and tissue sample. The specimen preparation device includes a plurality of elongated magnets movable from a first positon below the test tube bases to a second position adjacent the test tube lower cylindrical portions and a magnetic array mounting bracket for securing the elongated magnets parallel with one another. The specimen preparation device includes a plurality of elongated stop fins, each fin positioned to support one of the rows of test tubes by the test tube semispherical base. The fins extend parallel with the elongated magnets. For clarity, Fig. 10 shows only the first three rows of test tubes and only one of the plurality of magnets in a second position within the magnetic array chamber. Fig. 1 1 A shows the exploded view of three magnets mounted on the magnetic array mounting bracket and the test tube support plate having elongated openings for the magnets to move through when the magnets move from the first position to the second position. Fig. 1 1 B shows the magnetic array mounting bracket and the test tube support plate with the magnets in the second position and also shows the positon of test tubes (not shown) between the magnets with arrow Z when the magnets are in the second position. The magnetic array mounting bracket in Figs. 1 Ί A and 1 1 B only show three of the magnets for clarity, but the pictured embodiment includes eleven magnets in its normal operating environment. A mounting bracket may be manufactured to include any number of magnets. A heating block or nichrome wire heating element is positioned inside a rear duct and at least one fan is positioned in the front duct for circulating air through the ducts, allowing the heated air passed along a fan baffle and along the elongated stop fins and test tubes. The specimen preparation device includes a diffusion filter disposed adjacent the fan baffles for uniformly distributing the heat to the test tube array. A test tube support plate shown in Fig. 1 1 A has a plurality of parallel elongated slots, the slots sized to allow the elongated magnets to pass therethrough, the test tube support plate securing the elongated stop fins parallel with one another. The specimen preparation device includes a shaker assembly for moving the test tube support plate in a circular motion, the shaker assembly including a shaker bearing, shaker shaft, shaker pulley, shaker belt and shaker motor.

Figure 12 shows one of the plurality of elongated test tube stops. The stop includes a flat portion which may include slots for providing more even air circulation. The stop includes a mounting tab on each end for mounting the stop to the test tube support plate. A method of rinsing specimen samples using a specimen sample rinsing device may include providing the test tube support plate for securing a plurality of test tubes in a vertical position, the test tubes being in an array of parallel rows and providing a plurality of elongated magnets movable from a first positon, the elongated magnets horizontally between the rows of test tubes and vertically below level of the test tube bottoms. The magnets are movable to a second position adjacent the test tube cylindrical sides wherein the magnets are disposed on each side of the test tube and the magnet forces the iron filings and corresponding tissue sample against the sides of the test tube, allowing for the cleansing solution to be added and removed without removing the iron filings and tissue sample. The method includes adding the specimen sample and iron filings to at least one of the test tubes, moving the elongated magnets to the second position and adding a rinsing solution to the test tubes. The method includes removing the rising solution from the test tubes and repeating the steps of adding and removing the rinsing or cleansing solution until the sample is sufficiently clean. The elongated magnets may then be moved to the first position. At any time during the process, the test tubes may be shaken or vortexed with the elongated magnets in the first or second position in order to more thoroughly mix the cleaning solution with the specimen and iron filings. A similar method for rinsing specimen samples uses a specimen sample rinsing device comprising a test tube support plate for securing a plurality of test tubes in a vertical position. The test tubes may be in an array of parallel rows, the test tubes including a cylindrical wall and a semispherical base at the bottom end of the cylindrical wall. A plurality of elongated magnets are movable from a first positon wherein the elongated magnets are disposed horizontally between the rows of test tubes and vertically below level of the test tube bottoms to a second position wherein the magnets are disposed adjacent the test tube cylindrical sides. The method includes adding the specimen sample and iron filings to at least one of the test tubes and moving the elongated magnets from the first position to the second position wherein the magnets positioned on each side of the test tube force the iron filings and corresponding tissue sample against the sides of the test tube. The method includes adding a rinsing or cleansing solution to the at least one test tube wherein the force of the magnets allow for the cleansing solution to be added and removed without the removing the iron filings and tissue sample and removing the rising or cleansing solution from the at least one test tube. The steps of adding and removing the rinsing or cleansing may be repeated as many times as necessary to sufficiently clean the tissue sample. The elongated magnets may then be moved to the first position to allow the test tube tray to be removed for analysis.

Fig. 1 7 is a front view of the deflector system for selecting the heating or cooling mode. Fig. 18A is a left side view of the specimen preparation device with the deflectors in the heating mode and Fig. 18B is a left side view of the specimen preparation device with the deflectors in the cooling mode. Both Figs. 18A and 18B have the duct front cover, deflector gasket and the deflector end plate removed in order to show the position of the deflector in each mode. In the heating positon, the deflector plates adjacent both the inlet and outlet vents are positioned to cover the vents to prevent air from entering or exiting the device. In the heating mode, the air is circulated through the ducts, through the heating element and across the test tubes in order to raise or maintain the warmer temperature during the heating cycle. In the cooling mode shown in Fig. 18B, the deflectors are positioned about 90° from the heating position so that the lower duct is blocked and fresh air is forced through the side ducts and across the test tubes in order to cool the test tubes.

The following describes a method for controlling temperature of a specimen sample on a specimen preparation device. A sample of the ducts described is best seen in Fig. 1 B where the rear duct which has a rear duct cover and a rear duct side cover. A heater may be disposed within the duct cavity and may form a portion of the return duct. On a front portion of the specimen preparation device shown in Figs. 18A and 18B (duct panels removed) an inlet is disposed adjacent the front deflector which controls air through the inlet. The specimen preparation device has a forced air flow duct including a fan for forcing air through the system. The specimen preparation device includes a specimen air flow duct containing a specimen to be heated or cooled and a return air flow duct having a heater. The controllable inlet allows ambient air to enter the forced air flow duct and an controllable outlet allows air to exit the specimen preparation device. The method includes using the fan to flow air through the specimen air flow duct optially using heating or cooling through the specimen air flow duct and after air passes through the specimen, optionally flowing air through the return air duct to reenter the forced air flow duct. Optionally ambient air may be brought in to pass through the specimen air flow duct; and after passing air through the specimen air flow duct, air flows out of the outlet without the air flowing to the air flow inlet.

Figs. 13 - 18 show enlarged views of various portions of the specimen preparation device for clarity. Fig. 1 3 shows the test tubes resting on the elongated test tube stops. Fig. 14 shows detail on the test tubes, test tube units which is a row of test tubes, rack top and test tube support plate to which are mounted the elongated test tube stops. The rear tube deflector in Fig. 1 5 provides for increased temperature consistency across the test tubes and test tube stops. Fig. 1 6 shows details of the fan mounting brackets front, rear and side ducts and shaker components. Fig. 1 7 shows the configuration of the duct cover, gasket and deflector end plate. Fig. 18 is a side elevational view of the specimen preparation device with the main side panels removed and the front and rear ducts removed to show the deflector gaskets and the upper fan deflectors. Figs. 19 and 20 show the exploded view of the components within the magnetic array chamber. The entire chamber and contents may be shaken when the specimen preparation device is put into the vortexing mode.

While the present invention has been particularly described, in conjunction with a specific preferred embodiment, it is evident that many alternatives, modifications and variations will be apparent to those skilled in the art in light of the foregoing

description. It is therefore contemplated that the appended claims will embrace any such alternatives, modifications and variations as falling within the true scope and spirit of the present invention.

The specimen preparation device may use the vortexing system, magnetic separation system, and heating/cooling system in any combination to provide a variety of preparation sequences depending on the tests being performed on the sample. The samples may include body fluids, tissue or other samples for preparation.

Claims

1 . A specimen preparation device comprising: a device frame; a test tube support plate for securing a plurality of test tubes in a vertical position, the test tubes being in an array of parallel rows, the test tubes including a cylindrical wall and a base at the bottom end of the cylindrical wall; a plurality of elongated magnets movable from a first positon below the test tube bases to a second position adjacent the test tube lower cylindrical portions; a magnetic array mounting bracket for securing the elongated magnets parallel with one another; a plurality of elongated stop fins, each fin positioned to support one of the rows of test tubes by the test tube base, the fins extending parallel with the elongated magnets; a specimen air flow duct for providing a passage of air around or through the specimen, a return air duct including a heating element positioned inside the return air duct; a forced air flow duct having at least one fan positioned inside forced air flow duct, the at least one fan positioned adjacent a plurality of fan baffles for circulating air through and around the specimen air flow duct and the return flow air duct so that optionally heated air passes along the fan baffle and along the elongated stop fins and test tubes; a diffusion filter disposed adjacent the fan baffles for uniformly distributing the heat to the test tube array; a test tube support plate having a plurality of parallel elongated slots, the slots sized to allow the elongated magnets to pass therethrough, the test tube support plate securing the elongated stop fins parallel with one another; a shaker assembly for moving the test tube support plate in a circular motion, the shaker assembly including a shaker bearing, shaker shaft, shaker pulley, shaker belt and shaker motor; an inlet and outlet deflector movable to a first position wherein heated air is circulated through the device without the introduction of additional air to a second position wherein air is drawn in through the inlet duct, moved across the test tubes and is forced out through the outlet duct, allowing the test tubes to cool; wherein a volume of air may be circulated along the fins, heating the test tubes to a specified temperature, maintaining the specified temperature, cooling the volume of air and then repeating the heating and cooling process for a specified number of cycles; and wherein the test tubes may be shaken to mix contents of the test tube.

2. The specimen preparation device of claim 1 wherein the tissue samples within the test tubes are combined with iron filings or iron powder, the tissue sample being attracted to the iron filings during the shaking process whereby the iron filings are attracted to the sides of the test tube when the magnets are moved to the second positon and aspiration of the fluid and tissue in the test tube may occur by removing the fluid, adding a cleaning fluid to the test tube and with or without shaking the tube, removing the cleaning solution to leave substantially only the tissue sample attached to the iron filings.

3. The specimen preparation device of claim 1 wherein the plurality of test tubes may be a plurality of test tube units, each test tube unit being a row of test tubes permanently or removable secured in an elongated tube holder, the elongated tube holder securing a row of test tubes.

4. The apparatus of claim 1 where one or more of the following functions may be performed: vortexing the test tubes; moving the magnets adjacent the test tubes wherein magnetic filings in the test tube are attracted to the side of the test tube securing the iron filings against the wall or bottom of the test tube whereby a cleaning solution may be added to the test tube and shaken before or after the application of the magnets and the cleaning solution may be removed while the iron filings are secured to the wall or bottom of the test tube; heating the test tubes to a specified temperature; cooling the test tubes to a specified temperature

5. A specimen sample rinsing device for rinsing a plurality of samples combined with iron filings to which the samples are attracted to, the device comprising: a test tube support plate for securing a plurality of test tubes in a vertical position, the test tubes being in an array of parallel rows, the test tubes including a cylindrical wall and a semispherical base at the bottom end of the cylindrical wall; a plurality of elongated magnets movable from a first positon, the elongated magnets horizontally between the rows of test tubes and vertically below level of the test tube bottoms to a second position, the magnets adjacent the test tube cylindrical sides wherein the magnets positioned on each side of the test tube forces the iron filings and corresponding tissue sample to be forged against the sides of the test tube and allowing for the cleansing solution to be added and removed without removing the iron filings and tissue sample.

6. A method for controlling temperature of a specimen sample on a specimen preparation device comprising: providing a forced air flow duct including a fan; providing a specimen air flow duct containing a specimen to be heated or cooled; providing a return air flow duct having a heater; providing a controllable inlet for allowing ambient air to enter the forced air flow duct; providing an outlet for allowing air to exit the specimen preparation device; using the fan, flowing air through the specimen air flow duct optionally using heating or cooling through the specimen air flow duct; after air passes through the specimen, optionally flowing air through the return air duct to reenter the forced air flow duct; optionally bringing ambient air in through the inlet to pass through the specimen air flow duct; and after passing air through the specimen air flow optionally flowing air out of the outlet without the air flowing back to the specimen air flow duct.

7. A method of rinsing specimen samples using a specimen sample rinsing device comprising: providing a test tube support plate for securing a plurality of test tubes in a vertical position, the test tubes being in an array of parallel rows, the test tubes including a cylindrical wall and a semispherical base at the bottom end of the cylindrical wall; providing a plurality of elongated magnets movable from a first positon wherein the elongated magnets are disposed horizontally between the rows of test tubes and vertically below level of the test tube bottoms to a second position wherein the magnets are disposed adjacent the test tube cylindrical sides; adding the specimen sample and iron filings to at least one of the test tubes; moving the elongated magnets from the first position to the second position wherein the magnets positioned on each side of the test tube force the iron filings and corresponding tissue sample against the sides of the test tube; adding a rinsing or cleansing solution to the at least one test tube wherein the force of the magnets allow for the cleansing solution to be added and removed without the removing the iron filings and tissue sample; removing the rising or cleansing solution from the at least one test tube; repeating the steps of adding and removing the rinsing or cleansing solution for a desired number or cycles; moving the elongated magnets to the first position.

8. A method for preparing a specimen for analysis including using any combination of the methods in claims 6 or 7.

9. A method for preparing a specimen for analysis including using any combination of the methods in claims 6 or 7 and including shaking the test tubes between any of the steps.