JP2012177663A - Pre-freezer - Google Patents

Abstract

An object of the present invention is to provide a pre-freezer capable of easily and reliably pre-freezing an object to be frozen using various commercially available low-temperature liquefied gas containers.
A preliminary freezer 16 includes a bottomed cylindrical freezer body 17 inserted into a stopper portion of a cryogenic liquefied gas container, a preliminary freezing chamber 18 formed inside the freezer body, A heat induction shaft 19 that thermally penetrates the bottom of the freezer body and thermally couples the inside of the cryogenic liquefied gas container and the inside of the preliminary freezing chamber, and the sample container 24 enclosing the object to be frozen inside the preliminary freezing chamber. A sample container holding member 20 that is held and thermally coupled to the heat induction shaft, a lid member 21 that is attached to the upper opening of the preliminary freezing chamber, and a holding portion for holding the freezer body in the plug portion ( And a collar portion 22).
[Selection] Figure 1

Description

  The present invention relates to a pre-freezer, and in particular, pre-freezing for pre-freezing a biological sample or sample used for basic research such as biomedical, medical, pharmaceutical, etc. using a low temperature liquefied gas container by a slow freezing method. Related to the vessel.

  In general, when cryopreserving a biological sample, the biological sample is slowly pre-frozen to a certain temperature by a slow freezing method, and then immersed in cryogenic liquid nitrogen contained in a cryopreservation container and frozen. I try to save it. Conventionally, pre-freezing by the slow freezing method is a method in which a biological sample is enclosed in a sample container such as a microtube with a cap, a vial, an ampoule, or a bag together with a freezing medium, and is controlled at a rate of about −1 ° C./min while being controlled by a program freezer. The temperature is lowered and pre-frozen, or the biological sample is housed in a heat insulating case together with a freezing medium, and kept in a constant temperature refrigerator at about −80 ° C. for a predetermined time to be pre-frozen. In addition, as a method that does not use equipment such as a program freezer or a constant temperature refrigerator for pre-freezing, a freezer module that can pass through the lid of a cryocontainer containing biological samples is placed, and the operation of the cooling fan is controlled by the temperature control mechanism A method of pre-freezing a biological sample is known (for example, see Patent Document 1).

  On the other hand, as a cryopreservation container for storing the sample in a frozen state for a long period of time, a heat insulating container in which a heat insulating layer is formed between an outer tank and an inner tank is used, and in the upper part of the inner tank, A neck part with a smaller diameter than the inner tank is connected to the upper end of the neck part, and the upper part of the suspension part for holding the cryopreserved sample in a suspended state in the heat insulating container is suspended from the upper end opening part of the neck part. There is known a structure in which a locking member is provided and a heat insulating container including the locking member is covered with a cap (see, for example, Patent Document 2).

Japanese translation of PCT publication No. 2002-511826 JP 2007-271279 A

  However, in the case of using the above-described program freezer or constant temperature cold storage, if a power failure occurs or liquid nitrogen runs out, there is a possibility that the biological sample may be frozen poorly. In addition, liquid nitrogen is sprayed directly onto the microtube containing the biological sample in the preliminary freezing using the program freezer. Therefore, if there is any contamination in the liquid nitrogen or liquid nitrogen piping, the germ will enter through the gap in the cap of the microtube. There was a risk of contamination of the biological sample.

  Furthermore, in the method of Patent Document 1, although there is no possibility that various bacteria are mixed in the biological sample, the structure of the freezer module, the storage unit, and the temperature control mechanism is complicated, and the apparatus is increased in size and cost. In addition, the cryopreservation container of Patent Document 2 has no fear of freezing failure due to the occurrence of a power failure or the occurrence of liquid nitrogen breakage, and can prevent contamination by germs, but is intended for cryopreservation. Therefore, preliminary freezing could not be performed using this cryopreservation container.

  Therefore, the present invention has an object to provide a pre-freezer capable of easily and surely pre-freezing the object to be frozen by the slow freezing method of the object to be frozen using various commercially available low-temperature liquefied gas containers. Yes.

  In order to achieve the above object, the pre-freezer of the present invention is a pre-freezer that pre-freezes by a slow freezing method by being detachably attached to a cap portion provided in a low-temperature liquefied gas container that contains a low-temperature liquefied gas. A freezer main body comprising a bottomed cylindrical heat insulating material inserted into the plug portion, and having a preliminary freezing chamber therein; and the cryogenic liquefied gas passing through the bottom of the freezer body A heat induction shaft that thermally couples the inside of the container and the inside of the preliminary freezing chamber, and a sample container that encloses an object to be frozen is held inside the preliminary freezing chamber and is thermally coupled to the heat induction shaft. The sample container holding member, a lid member attached to the upper opening of the preliminary freezing chamber, and a holding part for holding the freezer main body in the plug part are provided.

  Furthermore, in the preliminary freezer of the present invention, the heat induction shaft includes a support member that supports the sample container holding member in the preliminary freezing chamber, and the support position of the sample container holding member is in the axial direction of the heat induction shaft. It is formed to be adjustable, heat transfer fins project from the outer surface of the tip portion inserted into the low-temperature liquefied gas container of the heat induction shaft, and the heat induction shaft A plurality of shaft members are connected in the axial direction of the shaft, and the cryogenic liquefied gas container is a liquid nitrogen type cryopreservation container.

  According to the preliminary freezer of the present invention, by using a freezer main body having a shape corresponding to the plug portion of various commercially available low-temperature liquefied gas containers, various kinds of samples such as biological samples can be obtained using various low-temperature liquefied gas containers. The frozen material can be frozen slowly. In addition, liquid nitrogen does not easily run out of power without being affected by a power failure, and there is no risk of invasion of germs. In particular, since the structure is simple, the manufacturing cost is low, and the handling is easy. Therefore, the cost required for the preliminary freezing by the slow freezing method can be reduced.

It is a partial cross section front view of the preliminary freezer which shows one example of the present invention. It is sectional drawing of the cryopreservation container which shows one use condition of a preliminary freezer.

  First, as shown in FIG. 2, a liquid nitrogen cryopreservation container 11 as a low-temperature liquefied gas container used when pre-freezing an object to be frozen by the slow freezing method in the preliminary freezer of the present invention is an outer tank 12. The inner tub 13 is composed of a cylindrical container main body 13a and a conical portion whose upper diameter is reduced on the upper part of the container main body 13a. A neck portion (plug portion) 13c provided continuously through 13b is integrally formed. The outer tub 12 is integrally formed with a cylindrical portion 12a that accommodates the container main body 13a via a heat insulating layer and a reduced diameter portion 12b whose upper diameter is reduced to accommodate the plug portion 13c. The upper end portion of the portion 12b and the upper end portion of the plug portion 13c are welded to integrally form the outer tub 12 and the inner tub 13.

  In addition, the cryopreserved sample is held in a suspended state in the inner tank 13 in the upper end opening of the plug portion 13c when performing long-term cryopreservation of the sample, which is the intended use of the cryopreservation container 11. A ring-shaped locking member 15 is provided for hooking an upper portion of a suspension member (not shown). Further, an outer lid 14a that covers the upper portion of the plug portion 13c is attached on the locking member 15, and a handle 14b that is used when carrying the cryopreservation container 11 is provided on the outer peripheral side of the upper surface.

  A pre-freezer 16 for pre-freezing a sample by the slow freezing method using the cryopreservation container 11 formed in this way is inserted into the mouth plug part 13c as shown in FIG. A freezer body 17 formed in a bottomed cylindrical shape, a preliminary freezing chamber 18 provided in an open state inside the freezer body 17, and a through hole 17 a provided in the bottom of the freezer body 17. A heat induction shaft 19 that thermally couples the inside of the preliminary freezing chamber 18 and the inside of the heat insulating container 14, the sample container holding member 20 that holds the sample container inside the preliminary freezing chamber 18, and the preliminary freezing A lid member 21 attached to the upper opening of the chamber 18 and a flange portion 22 protruding from the outer periphery of the upper portion of the freezer body 17 as a holding portion for holding the freezer body 17 in a predetermined position of the plug portion 13c. I have.

  The freezer body 17 is formed of a heat insulating material such as foamed vinyl chloride, foamed urethane, or foamed silicon, and the outer diameter of the freezer body 17 is made smaller than the inner diameter of the plug portion 13c or the freezer body 17 is made. It is formed so that the gas vaporized in the heat insulation container 14 can be discharged | emitted outside by providing the ditch | groove of an axial direction in the outer peripheral surface. Further, the flange 22 can be formed integrally with the freezer body 17. On the other hand, the heat induction shaft 19 and the sample container holding member 20 are formed of a metal such as aluminum or copper having good heat conductivity.

  The heat induction shaft 19 is inserted into the lower heat induction shaft 19a fitted in the through hole 17a communicating the inside of the preliminary freezing chamber 18 and the inside of the heat insulating container 14, and inserted into the upper end of the lower heat induction shaft 19a. The upper heat induction shaft 19b is divided into a thermally coupled upper heat induction shaft 19b. A suspension rod 23 formed of a synthetic resin having low heat conductivity is detachably attached to the upper portion of the upper heat induction shaft 19b. Yes.

  The sample container holding member 20 has a cylindrical shape provided with a through hole 20a through which the upper heat induction shaft 19b is inserted at the center, and a microtube in which an object to be frozen is sealed around the through hole 20a. A plurality of receiving holes 20b for holding a sample container 24 such as the like are provided at equal intervals in the circumferential direction. The sample container holding member 20 is supported at a predetermined position in the preliminary freezing chamber 18 by a support member 25 made of a ring-shaped elastic member such as silicon rubber attached to the upper heat induction shaft 19b. On the outer periphery of the upper heat induction shaft 19b, a plurality of circumferential grooves 25a for mounting the support member 25 are provided in the vertical direction (axial direction), and by selecting the circumferential groove 25a for mounting the support member 25, The support height of the sample container holding member 20 in the preliminary freezing chamber 18 can be adjusted.

  The upper part of the suspension rod 23 passes through a through hole 21a provided in the center of the lid member 21, and further passes through a through hole 14c formed in the outer lid 14a so as to protrude above the outer lid 14a. A spherical knob 23a is provided at the upper end of the suspension bar 23 protruding from the top. The through hole 14c of the outer lid 14a may have a diameter that allows the knob 23a to be inserted.

  Further, the lower heat induction shaft 19a is positioned on the upper side of the lower heat induction shaft 19a by contacting the bottom surface of the freezer body 17 and restricting the amount of the lower heat induction shaft 19a inserted into the through hole 17a. A flange portion 19c is provided, and a plurality of heat transfer fins 19d are projected radially at equal intervals around the lower heat induction shaft 19a at the lower end portion of the lower heat induction shaft 19a.

  When the object to be frozen, for example, a biological sample, is pre-frozen by the slow freezing method using the cryopreservation container 11 and the pre-freezer 16, the upper end of the lower heat induction shaft 19a is inserted into the through hole 17a of the freezer body 17. The lower heat induction shaft 19a is inserted from the plug portion 13c into the heat insulating container 14 of the cryopreservation container 11 storing a predetermined amount of low-temperature liquefied gas, usually liquid nitrogen. The freezer body 17 is mounted at a predetermined position in the plug portion 13c by bringing the flange portion 22 of the freezer body 17 into contact with the upper surface of the locking member 15.

  On the other hand, the upper heat induction shaft 19b is inserted from below into the through hole 20a of the sample container holding member 20 that holds the sample container 24 enclosing the biological sample in the accommodation hole 20a, and is supported by the support member 25 attached to the predetermined circumferential groove 25a. The sample container holding member 20 is supported, the sample container holding member 20 is inserted into the preliminary freezing chamber 18 of the freezer body 17, and the lower end portion of the upper heat induction shaft 19b is opened at the upper end portion of the lower heat induction shaft 19a. To the lower heat induction shaft 19a and the upper heat induction shaft 19b. Further, the lid member 21 having the suspension rod 23 inserted into the through hole 21 a is attached to the upper opening of the preliminary freezing chamber 18.

  Thereby, the cold heat of liquid nitrogen is transmitted to the sample container 24 through the heat transfer fins 19d, the lower heat induction shaft 19a, the upper heat induction shaft 19b, and the sample container holding member 20 each formed of a material having good heat transfer properties. Then, the biological sample in the sample container 24 is slowly cooled. At this time, the liquid surface height of the liquid nitrogen in the heat insulating container 14, the positions of the heat transfer fins 19d, the lengths of the lower heat induction shaft 19a and the upper heat induction shaft 19b, the lower heat induction shaft 19a and the upper heat induction shaft 19b The thermal coupling state of the upper heat induction shaft 19b and the sample container holding member 20, the position of the sample container holding member 20 in the preliminary freezing chamber 18, the accommodation hole 20a of the sample container holding member 20 and the sample container By appropriately setting various conditions such as the thermal coupling state with 24, the number of sample containers 24, and the state and amount of the biological sample, the cooling rate of the biological sample is set to an appropriate range, for example, about -1 ° C / min. The cooling rate can be set.

  For example, the cooling rate can be increased by lowering the temperature of the heat transfer fin 19d by immersing the heat transfer fin 19d in liquid nitrogen, and conversely, the heat transfer fin 19d is higher than the liquid nitrogen level. The cooling rate can be reduced by setting the position. Further, since the temperature in the preliminary freezing chamber 18 becomes relatively higher as it is closer to the lid member 21, the sample is placed on the bottom side of the preliminary freezing chamber 18 by setting the circumferential groove 25 a for mounting the support member 25 on the upper side. The cooling rate can be reduced by increasing the temperature during preliminary freezing compared to when the container holding member 20 is positioned.

  In this way, various samples can be pre-frozen by the slow freezing method by simply using a commercially available ordinary cryopreservation container 11 and attaching the pre-freezer 16 to the stopper 13c of the cryopreservation container 11. Since the only cooling source for pre-freezing the object to be frozen is liquid nitrogen in the cryopreservation container 11, there is no problem even if a power failure occurs during the pre-freezing process, and the consumption of liquid nitrogen is also small. The cost of liquid nitrogen can be reduced without running out of liquid. Furthermore, since the sample container 24 is in a state of being enclosed in the preliminary freezing chamber 18, various germs do not enter from the outside.

  Therefore, it is possible to greatly reduce the cost of pre-freezing biological objects such as biological samples by the slow freezing method, and to easily and surely carry out the pre-freezing process. Since no equipment is required, the cost required for the preliminary freezing process can be greatly reduced and the processing efficiency can be greatly improved. Further, by using the cryopreservation container used for cryopreservation of the sample for pre-freezing, the process from the pre-freezing to the cryopreservation can be performed with only one cryopreservation container, so that the number of equipment used can be reduced. On the other hand, not only the cryopreservation container but also other members other than the freezer body 17 by appropriately forming the freezer body 17 of the preliminary freezer 16 in a shape corresponding to the shape of the plug portion of various cryogenic liquefied gas containers. Can be used in common, not only the cryopreservation container as described above, but also various cryogenic liquefied gas containers can be used to pre-freeze various objects to be frozen using the pre-freezer 16. .

  Sample containers that have been pre-frozen after a preset time have passed can be removed from the pre-freezer and stored in the same manner as before. It can also be used as a container. In addition, the lower heat induction shaft and the upper heat induction shaft can be formed as a single continuous heat induction shaft, and the heat induction shaft can be extended and contracted. Further, the position of the sample container holding member relative to the heat induction shaft may be fixed, and the heat transfer fins may be omitted. Moreover, each material can be selected in consideration of heat conductivity, heat insulation, strength, moldability, and the like, and is not particularly limited.

  DESCRIPTION OF SYMBOLS 11 ... Cryopreservation container, 12 ... Outer tank, 12a ... Cylindrical part, 12b ... Reduced diameter part, 13 ... Inner tank, 13a ... Container main body, 13b ... Conical part, 13c ... Mouth plug part, 14 ... Thermal insulation container, 14a ... Outer lid, 14b ... handle, 14c ... through hole, 15 ... locking member, 16 ... pre-freezer, 17 ... freezer body, 17a ... through-hole, 18 ... pre-freezing chamber, 19 ... heat induction shaft, 19a ... bottom Heat induction shaft, 19b ... Upper heat induction shaft, 19c ... Gutter, 19d ... Heat transfer fin, 20 ... Sample container holding member, 20a ... Through hole, 20b ... Housing hole, 21 ... Cover member, 21a ... Through hole, 22 ... Hut, 23 ... Hanging rod, 23a ... Pick part, 24 ... Sample container, 25 ... Support member, 25a ... Circumferential groove

Claims (5)

A pre-freezer that is detachably attached to a stopper provided in a low-temperature liquefied gas container that contains a low-temperature liquefied gas and performs preliminary freezing by a slow freezing method, and is inserted into the stopper. A freezer body made of a bottom cylindrical heat insulating material and having a preliminary freezing chamber inside, and thermally passing between the inside of the cryogenic liquefied gas container and the inside of the preliminary freezing chamber through the bottom of the freezing body A heat induction shaft to be coupled, a sample container enclosing the object to be frozen is held in the preliminary freezing chamber, a sample container holding member thermally coupled to the heat induction shaft, and an upper opening of the preliminary freezing chamber A pre-freezer comprising a lid member to be mounted and a holding part for holding the freezer main body in the plug part. 2. The heat induction shaft includes a support member that supports the sample container holding member in the preliminary freezing chamber, and a support position of the sample container holding member is formed so as to be adjustable in an axial direction of the heat induction shaft. The pre-freezer described. The preliminary freezer according to claim 1 or 2, wherein the heat induction shaft is provided with a heat transfer fin projecting from an outer surface of a tip portion inserted into the low temperature liquefied gas container. The pre-freezer according to any one of claims 1 to 3, wherein the heat induction shaft is formed by connecting a plurality of shaft members in an axial direction of the heat induction shaft. The preliminary freezer according to any one of claims 1 to 4, wherein the low-temperature liquefied gas container is a liquid nitrogen type cryopreservation container.

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