WO2022019737A1 - Cryostat automatique portatif - Google Patents

Cryostat automatique portatif Download PDF

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Publication number
WO2022019737A1
WO2022019737A1 PCT/MX2020/000021 MX2020000021W WO2022019737A1 WO 2022019737 A1 WO2022019737 A1 WO 2022019737A1 MX 2020000021 W MX2020000021 W MX 2020000021W WO 2022019737 A1 WO2022019737 A1 WO 2022019737A1
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WO
WIPO (PCT)
Prior art keywords
cryostat
shaft
base
motor
collection module
Prior art date
Application number
PCT/MX2020/000021
Other languages
English (en)
Spanish (es)
Inventor
Jesús Raúl BELTRÁN RAMÍREZ
Original Assignee
Beltran Ramirez Jesus Raul
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Beltran Ramirez Jesus Raul filed Critical Beltran Ramirez Jesus Raul
Publication of WO2022019737A1 publication Critical patent/WO2022019737A1/fr

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/02—Devices for withdrawing samples
    • G01N1/04—Devices for withdrawing samples in the solid state, e.g. by cutting
    • G01N1/06—Devices for withdrawing samples in the solid state, e.g. by cutting providing a thin slice, e.g. microtome
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/42—Low-temperature sample treatment, e.g. cryofixation

Definitions

  • the present invention relates to the technical field of mechanics, electronics and obtaining histological samples, since it provides a portable automatic cryoatate. BACKGROUND OF THE INVENTION
  • a cryoatate (from cryo "cold” and stat “stable”) is a device used to maintain cryogenically low temperatures of samples or devices mounted within the cryoatate. Low temperatures can be maintained within a cryostat using various methods of refrigeration, most commonly using a cryogenic fluid bath such as liquid helium. Therefore, it is usually mounted in a vessel, similar in construction to a vacuum flask or Dewar. Cryoatates have numerous applications in science, engineering, and medicine.
  • the cryoatate is used in the processing of samples for diagnosis or histological identification. It is mainly used to freeze and obtain sections of tissues or frozen samples that have not been previously fixed by chemical methods, to prevent them from losing some important characteristics necessary for observation.
  • Frozen sectioning is the method for preparing a selected portion of tissue for pathology purposes.
  • the tissue is moistened and, with or without fixation, snap frozen and sectioned with a microtome in a cryostat.
  • the technique is very fast and allows examination of the sample obtained during the surgical intervention, whose evolution will depend on the report issued by the pathologist.
  • Frozen sections are essential for certain staining methods, for example, the demonstration of fat by the red or oil method, and some silver impregnation methods for the study of the central nervous system. Frozen sections are also essential for immediate examination during procedures. In frozen sectioning, the tissue does not decrease in volume (except when cleared and mounted with xylene, etc.), whereas ordinary paraffin-mounted sections shrink by 10 to 15 percent.
  • the cryostat is used to obtain frozen sections approximately 10 to 30 ⁇ m thick.
  • the entire cutting system is enclosed in a refrigerated chamber whose temperature can be regulated, normally between -20 and -30 °C. Freezing is usually done on a platform inside the refrigerated chamber itself, which is at a lower temperature, around -50 °C.
  • the tissue can also be frozen externally as quickly as desired, for example, with liquid nitrogen, but it is convenient to place the sample in the cryostat chamber until its temperature is equal to its temperature, in order to obtain homogeneous sections.
  • the sample Before freezing, the sample is embedded in a medium that is liquid at room temperature and solid at shear temperature. Thus, we have our sample in a solid block, embedded but not included. This allows the sample to be manipulated and adhered to a sample holder support, which will be fixed to a shaft that advances on the blade.
  • the block preparation and cutting mechanism is similar to that of the rotary paraffin microtome.
  • the sections that are obtained are adhered by contact to slides with adhesive surfaces. The sections thaw quickly in this gluing process since the slides are at room temperature and once dry they can be processed for the techniques we want.
  • cryostats There are different types of cryostats which have been developed to improve cooling and cutting techniques.
  • CLOSED CYCLE CRYOSTATS They consist of a chamber through which cold helium vapor is pumped. An external mechanical cooler extracts the hot helium vapor, which is cooled and recycled. Closed-loop cryostats consume a relatively large amount of electrical power, but do not have to be refilled with helium and can run continuously for an indefinite period. Objects can be cooled by attaching to a metallic cooling plate inside a vacuum chamber that is in thermal contact with the helium vapor chamber.
  • CONTINUOUS FLOW CRYOSTATS They are cooled by liquid cryogens (helium or normally liquid nitrogen) from a storage flask. As the cryogen boils inside the cryostat, it is continually replenished by a constant flow from the storage flask. Temperature control of the sample within the cryostat is typically done by controlling the flow rate of cryogen into the cryostat in conjunction with a heating wire attached to a temperature control loop. The length of time that refrigeration can be maintained is controlled by the volume of cryogens available. Due to the shortage of liquid helium, Some laboratories have facilities to capture and recover the helium as it escapes from the cryostat, although these facilities are also expensive to operate.
  • liquid cryogens helium or normally liquid nitrogen
  • BATH CRYOSTATS They are similar in construction to vacuum flasks filled with liquid helium. A cooling plate is placed in thermal contact with the liquid helium bath. Liquid helium can be replenished, as it boils away, at intervals between a few hours and several months, depending on the volume and construction of the cryostat. The rate of evaporation is minimized by shielding the bath, either with cold helium vapor, or a vacuum shield with walls constructed from a material called super insulation. Helium vapor boiling away from the bath is very effectively cooled by heat shields around the outside of the bath. In older designs there may be an additional bath of liquid nitrogen, or several concentric layers of shielding, with gradually increasing temperatures. However, the invention of super insulating materials has made this technology obsolete.
  • MULTI-STAGE CRYOSTATS In order to achieve a lower temperature than liquid helium, additional colder stages can be added to the cryostat, where temperatures up to 1K can be achieved, by attaching the cooling plate to a 1K pot, which is a container of He-3, isotope that is connected to the vacuum pump. Temperatures down to 1MK can also be achieved using a dilution cooler or dry dilution cooler, typically in addition to the main stage and 1K pot, lower temperatures than magnetic cooling can achieve.
  • ULTRA-CRYOSTAT This device is used to obtain ultra-thin sections, of the order of tens of nanometers, for observation with the transmission electron microscope.
  • ACTUATOR which describes a mechanical drive for remotely applying force to a cryogenically cooled device, having a mechanical actuator made up of multiple parts. During operation, the parts bear against each other to allow an actuating device to apply a force, and when not in use, the parts come apart.
  • the document cited above refers to a mechanism that allows force to be exerted remotely on a cryogenically frozen device, but does not show evidence of having cooling means integrated in said device, nor does it describe having a cutting mechanism and Automatic sample collection.
  • Figure 1 shows a perspective view of the transparency of the portable automatic cryostat.
  • Figure 2 shows a perspective view of the motor and the auger of the portable automatic cryostat.
  • Figure 3 shows a perspective view of the sample base of the portable automatic cryostat.
  • Figure 4 shows a bottom view of the sample base of the portable automatic cryostat.
  • Figure 5 shows a top perspective view of the portable automatic cryostat plate.
  • Figure 6 shows a top view of the portable automatic cryostat plate.
  • Figure 7 shows a side perspective view of the movement mechanism of the portable automatic cryostat.
  • Figure 8 shows a rear perspective view of the movement mechanism of the portable automatic cryostat.
  • Figure 9 shows a side view of the movement mechanism and the cutting module of the portable automatic cryostat.
  • Figure 10 shows a side perspective view of the cutting module of the portable automatic cryostat.
  • Figure 11 shows a side perspective view of the collection module of the portable automatic cryostat.
  • Figure 12 shows a perspective view of the support of the portable automatic cryostat.
  • Figure 13 shows a perspective view of the portable automatic cryostat knife holder.
  • the portable automatic cryostat is made up of a structure (1), which preferably it is cylindrical in shape, which is configured to house in the lower part, a motor base (2); a motor (3) is installed on the motor base (2) and a worm (4) is installed from one of its ends on the shaft of the motor (3).
  • a base for samples (5) preferably prismatic in shape, is installed on the auger (4), in the strange opposite of the motor (3), and is installed by means of a hole (6) which is threaded in its interior, as shown in figure 4, this configuration allows the rotation of the motor (3) to be transmitted to the base for samples (5) by means of the screw (4) and transform the rotation into linear movement so that said base for samples (5) can scroll vertically up or down;
  • a sample holder (not shown) is located on top of the sample base (5) which is configured to hold samples that are preferably in paraffin blocks.
  • At least one linear guide (7) is located on the outside of any of the sides of the base for samples (5) which is configured to assemble in at least one rail (8) that is inside the structure (1) aligned to the linear guide (7), this configuration allows that, when the base for samples (5) is moved up or down, it makes a vertical movement without lateral movements.
  • a cutting module (12) is installed on the periphery of the upper part of the plate (9) placed on one of the sides of the hole (10); a collection module (13) is installed on the plate (9) opposite the cutting module (12), on the opposite side of the hole (10).
  • the cutting module (12) and the collection module (13) are each made up of a movement mechanism (14), which has two shaft bases (15) installed in parallel; a shaft (16) is installed on top of the shaft bases (15), the lateral ends of said shaft (16) protrude from the shaft bases (15) on which an impeller (17) is installed which is configured to install a transmission gear (18) on its shaft; a rotation gear (19) is installed on the shaft (16) coupled with the transmission gear (18), this configuration allows the rotation of the impeller (17) to be transmitted to the rotation gear (19) and rotate the shaft ( 16); a preferably eccentric cam (20) is installed on each of the protruding ends of the shaft (16).
  • a support (21) of preferably rectangular prismatic shape is installed in the front part of each of the movement mechanisms (14), said support (21) has on its surface, two perforations (22) which are in parallel and cross the thickness of said support (21), and are configured to install some rods (23), where their ends protrude from both sides of said support (21);
  • a cam follower (24) is installed at the rear of the rods (23) and are configured to make contact with the smaller radius of the cam (20) and convert the rotary movement of the shaft (16) into linear movement, moving said rods (23) forward;
  • a preferably compression spring (25) is located at the rear of the rods (23) between the support (21) and the cam follower (24), this configuration allows the rods (23) to move backwards when the radius cam follower (20) stops making contact with cam follower (24).
  • a blade holder (26) is located on the rods (23), installed at the opposite end of the cam followers (24) of the cutting module (12), said blade holder (26) is configured to install a blade (27 ), this allows cuts to be made in a sample when the smaller radius of the cam (20) makes contact with the cam follower (24) and the rods (23) are moved forward.
  • a slide base (28) is located on the rods (23), installed at the opposite end of the cam followers (24) of the harvesting module (13), said slide base (28) is configured to install a slide (29), this allows to collect the samples of the cuts made by the blade (27) of the cutting module (12) when the smaller radius of the cam (20) makes contact with the cam follower (24) and the rods ( 23) are pushed forward.
  • the cutting module (12) and the collection module (13) are configured to be activated in a synchronized manner, in such a way that when the blade (27) cuts the sample, which falls on the slide base ( 28).
  • At least one cooling medium (30) which is preferably a peltier plate, is installed in the opening (11) of the plate (9), which is configured to maintain a lower outside temperature in the upper part of the structure (1).
  • a removable cover (31), preferably concave, is installed in the upper part of the structure (1) allowing a hermetic closure, said cover (31) has a concavity (33) in its circumference where the collection module is placed (13) and the movement mechanism (14) of said collection module (13) in such a way that they remain outside the lid (31), this configuration allows the sample found in the base for samples to be isolated from the outside ( 5), the cutting module (12), the slide (29), and at the same time maintain the temperature generated by the cooling medium (30); a sliding gate (not illustrated) is placed in the concavity (33) of the lid (31), which is configured to take the samples obtained by the collection module (13).
  • a duct (not illustrated) has one of its ends installed in the lower part of the cooling medium (30), and the opposite end is installed in the collection module (13) aligned with the slide (29), this configuration allows maintaining a point temperature control in said slide (29) maintaining a higher temperature than the one inside the lid (31) and being able to collect the samples obtained by the cutting module (12) with the slide (29) by thermal difference .
  • a switch (not illustrated) is installed on the outer periphery of the structure (1), which is configured to pass electrical energy from a power source. (not illustrated) to a control module (32) that is installed on the outer periphery of said structure (1), said control module (32) is configured to control the operating parameters of the portable automatic cryostat.
  • a potentiometer (not illustrated) is installed on the outer periphery of the structure (1), which is configured to program the advance of the motor (3) and define the cutting thickness of each of the samples found on the base. for samples (5).
  • a temperature control (not illustrated) is installed on the outer periphery of the frame (1), and is configured to adjust the temperature of the cooling medium (30).
  • a preferably tactile screen (not illustrated) is located outside the structure (1), which is configured to display the operating parameters, such as temperature, sample thickness, number of cuts to be made, among others.
  • An activation button (not illustrated) is located on the outer periphery of the structure (1) which is configured to initiate the operation of the portable automatic cryostat.
  • the motor (3) and drivers (16) can be an electric motor, a servo motor or a stepper motor.
  • the power source (not shown) can be a direct current cable, a battery, a portable charger and/or a combination of the above.
  • a paraffin sample is placed in the sample base (5), then a knife (27) is installed in the knife holder (26), then a slide (29) is placed in the base of slides (28), and finally the lid (31) is placed on the structure (1) to protect the sample, as well as the cutting module (12) and at the same time isolate from the outside to contain the temperature of said sample .
  • the switch (not illustrated) is activated to energize the control module (32) by means of the power source (not illustrated), the potentiometer (not illustrated) is adjusted to program the thickness of the samples by means of the advance of the motor (3), finally the temperature of the cooling medium is adjusted with the temperature control (not illustrated) and the configuration data is corroborated by means of the screen (not illustrated).
  • R EA presses the activation button (not illustrated) and the motor (3) rotates, transmitting the rotation through the auger (4) to move the base for samples (5) upwards, once the sample is positioned in front of the cutting module (12) and the collection (13), the drivers (17) are activated to move the rods (23) by means of the cams (20) and that both the knife (27) and the slide (29) approach the samples, once Once the cut has been made, the sample is deposited on the slide (29) and the rods (23) of the cutting module (12) and of the collection module (13) move back by means of the springs (25).

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  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Sampling And Sample Adjustment (AREA)
  • Automatic Analysis And Handling Materials Therefor (AREA)

Abstract

La présente invention concerne un cryostat automatique portatif qui comprend des moyens de réfrigération intégrés permettant de maintenir une température inférieure à la température extérieure, étant donné que, au moyen de son couvercle, l'intérieur du cryostat est isolé, en outre, les échantillons qui sont dans la base d'échantillons sont protégés d'une éventuelle contamination. Le cryostat de la présente invention présente un module de coupe et un module de prélèvement d'échantillons, lesquels fonctionnent simultanément de manière automatique et indépendante. Le cryostat automatique portatif présente un système de régulation de température ponctuelle, ce qui permet une variation de température dans le module de prélèvement d'échantillons différente à l'intérieur où sont effectuées les coupes, ce qui permet de recueillir les échantillons par différence thermique.
PCT/MX2020/000021 2020-07-20 2020-07-24 Cryostat automatique portatif WO2022019737A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
MX2020007712A MX2020007712A (es) 2020-07-20 2020-07-20 Criostato automatico portatil.
MXMX/A/2020/007712 2020-07-20

Publications (1)

Publication Number Publication Date
WO2022019737A1 true WO2022019737A1 (fr) 2022-01-27

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WO (1) WO2022019737A1 (fr)

Citations (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DE1236818B (de) * 1965-01-11 1967-03-16 Wkf Ges Fuer Elektrophysikalis Einrichtung zur Gefriertrocknung einer mittels eines Gefriermikrotoms hergestellten Stoffprobe
WO1987002130A1 (fr) * 1985-10-04 1987-04-09 Brian Laurence Jackson Ameliorations apportees a des microtomes cryostatiques
FR2705587A1 (fr) * 1993-05-25 1994-12-02 Tabone Herve Cryostat de microtomie, notamment pour travaux histologiques.
US5711200A (en) * 1994-09-30 1998-01-27 Leica Instruments Gmbh Cryostatic microtome
US5960640A (en) * 1996-09-30 1999-10-05 Microm Laborgerate Gmbh Cryostatic microtome
US20030167892A1 (en) * 2002-03-09 2003-09-11 Klaus Foerderer Feeding mechanism for a microtome
EP1811280A1 (fr) * 2006-01-18 2007-07-25 Seiko Instruments R&D Center Inc. Appareil de fabrication d'article tranché automatiquement et appareil de fabrication d'échantillon d'article tranché automatiquement
EP2607879A1 (fr) * 2010-08-18 2013-06-26 Sakura Seiki Co., Ltd. Microtome et cryostat
US20160123847A1 (en) * 2008-01-18 2016-05-05 Leica Mikrosysteme Gmbh Micromanipulator for a cryomicrotome
CN106353126A (zh) * 2016-07-30 2017-01-25 关慧娟 快速冷冻切片装置
CN206920145U (zh) * 2017-07-24 2018-01-23 李艳丽 一种病理研究专用自动冰冻切片机
CN208999118U (zh) * 2018-08-14 2019-06-18 上海菱蓝生物科技有限公司 恒冷病理切片机

Patent Citations (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DE1236818B (de) * 1965-01-11 1967-03-16 Wkf Ges Fuer Elektrophysikalis Einrichtung zur Gefriertrocknung einer mittels eines Gefriermikrotoms hergestellten Stoffprobe
WO1987002130A1 (fr) * 1985-10-04 1987-04-09 Brian Laurence Jackson Ameliorations apportees a des microtomes cryostatiques
FR2705587A1 (fr) * 1993-05-25 1994-12-02 Tabone Herve Cryostat de microtomie, notamment pour travaux histologiques.
US5711200A (en) * 1994-09-30 1998-01-27 Leica Instruments Gmbh Cryostatic microtome
US5960640A (en) * 1996-09-30 1999-10-05 Microm Laborgerate Gmbh Cryostatic microtome
US20030167892A1 (en) * 2002-03-09 2003-09-11 Klaus Foerderer Feeding mechanism for a microtome
EP1811280A1 (fr) * 2006-01-18 2007-07-25 Seiko Instruments R&D Center Inc. Appareil de fabrication d'article tranché automatiquement et appareil de fabrication d'échantillon d'article tranché automatiquement
US20160123847A1 (en) * 2008-01-18 2016-05-05 Leica Mikrosysteme Gmbh Micromanipulator for a cryomicrotome
EP2607879A1 (fr) * 2010-08-18 2013-06-26 Sakura Seiki Co., Ltd. Microtome et cryostat
CN106353126A (zh) * 2016-07-30 2017-01-25 关慧娟 快速冷冻切片装置
CN206920145U (zh) * 2017-07-24 2018-01-23 李艳丽 一种病理研究专用自动冰冻切片机
CN208999118U (zh) * 2018-08-14 2019-06-18 上海菱蓝生物科技有限公司 恒冷病理切片机

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