CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This invention claims priority of the German patent application 100 65 143.7 which is incorporated by reference herein.

FIELD OF THE INVENTION

[0002] The invention concerns a specimen holder for a high-pressure freezing device, according to the preamble of claim 1.

BACKGROUND OF THE INVENTION

[0003] For purposes of the invention, a “high-pressure freezing device” is understood to be a freezing system for rapid freezing (vitrification) of water-containing specimens under high pressure. Devices of this kind are described in DE Patent 18 06 741, EP 0 853 238 A1, and EP 0 637 741 A1.

[0004] A high-pressure freezing device of this kind is being successfully marketed by the Applicant under the name “Leica EM HPF,” and is depicted in the document “LEICA EM HPF, High Pressure Freezer, 1.K.-LEICA EM HPF-E-6/94, Juni 1994.”

[0005] The “Leica EM HPF” high-pressure freezing device makes it possible to vitrify conventional specimens under a pressure of approximately 2000 bar at a cooling rate of 10³-10⁵ K/s. With this known unit, the critical cooling phase from room temperature to −100° C. lasts approximately 10 ms at the surface of the specimen (cooling rate of 10⁴ K/s).

[0006] The cooling rate in the interior of the specimen depends exclusively on the physical properties of the specimen. In order to vitrify thicker biological or water-containing specimens, the specimen is exposed to high pressure.

[0007] Because of the elevation in pressure, a slower cooling rate is sufficient for vitrification. The cooling rate for vitrification of biological specimens is approximate 10⁵-10⁶ K/s at standard pressure, but at 2000 bar the cooling rate is only 10³-10⁴ K/s; in other words, under high pressure biological specimens can still be vitrified at cooling rates that are a hundred times lower.

[0008] Metal holders that comprise at least two shaped parts detachably joinable to one another are used for freezing the specimens, the shaped parts joined to one another forming a chamber for reception of the specimens. Specimen holders of this kind are depicted in the aforementioned document “LEICA EM HPF, High Pressure Freezer, 1.K.-LEICA EM HPF-E-6/94, Juni 1994” and in the document “Balzers, Hochdruck-Gefriermaschine [High-pressure freezing machine] HPM 010, Balzers Union Aktiengesellschaft, no publication date.”

[0009] In order to freeze the specimen, the specimen holder is clamped in the high-pressure freezing device in such a way that on the one hand specimen loss is prevented, and on the other hand a sufficient portion of the surface remains freely accessible for a cryogen, e.g. liquid nitrogen, to be sprayed onto it. An arrangement of this kind is depicted and described in unpublished DE 100 15 773.

[0010] From the known art of refrigeration and from the physical processes, it is also known that the cooling rate at the center of a biological specimen is determined only to a limited extent by the cooling rate at the surface. The essential feature in this context is that with rapid cooling rates at the surface, a kind of “saturation effect” occurs, whereas lower cooling rates at the surface inevitably lead to poor freezing results. It is therefore essential that the cooling rate achievable by a unit be transferred in as undiminished a fashion as possible by the specimen holding system onto the surface of the biological specimen.

SUMMARY OF THE INVENTION

[0011] It is therefore the object of the present invention to create a specimen holder for a high-pressure freezing device that transfers the achievable cooling rate in as undiminished a fashion as possible onto the surface of the specimen.

[0012] According to the present invention, this object is achieved by the features recited in the characterizing portion of claim 1. Advantageous developments of the invention are the subject matter of the dependent claims.

[0013] The invention is characterized in that one of the shaped parts is made of diamond or at least comprises a diamond. Diamond possesses not only extreme hardness but also the advantage of low specific heat and very good thermal conductivity.

[0014] In a further embodiment of the invention, at least one of the shaped parts, preferably the shaped part fabricated of diamond, is of disk-shaped configuration. A spacer ring fabricated from metal, which determines the depression necessary for specimen reception, is provided between the two shaped parts. Said spacer ring can also assume the sealing function in this context.

[0015] Provision can, of course, also be made for only one of the shaped parts to be fabricated of diamond or for the shaped part to comprise a mounted diamond. In this case the depression for specimen reception can be accomplished by the mount of the diamond or by a corresponding configuration of the other shaped part. A preferred exemplary embodiment is a specimen holder having one disk-shaped shaped part made of diamond, and one disk-shaped one made of metal. The metal shaped part has a circumferential shaped-on bead at the rim.

[0016] In a further embodiment of the specimen holder, the diamond or one of the shaped parts can be equipped with an orifice for the delivery of high pressure.

[0017] It has proven to be advantageous in the context of such specimen holders to use gold or aluminum or copper as the metal, and to utilize a commercially available polycrystalline CVD industrial diamond as the diamond.

[0018] In a further embodiment of the invention, both shaped parts are of planar configuration on their inwardly directed surfaces. A spacer running around the surfaces is arranged between those surfaces as a seal and in order to constitute the specimen receptacle. Differently dimensioned receptacles for the specimens can easily be produced by way of differently dimensioned spacers.