HPM 010 High Pressure Freezing Machine

The Proven High Pressure Freezing Machine

With over 100 installations world-wide, and more than 400 scientific publications to its credit, the HPM-010 is not only the pioneer of HP freezing, but the instrument of choice which allows freezing of aqueous samples up to 200µm thickness without visible ice crystal damage and without the use of cryo-protectants.

Features

• Freezing of specimens up to 200µm thickness and 2mm diameter without visible ice crystal damage
• No need for cryo-protectants
• Cryo-fixation of suspensions, monolayer cell cultures and tissue
• Short handling time before freezing
• Reproducible freezing
• Easy one button operation
• Large base of application know-how available

Features / Benefits

• Highest quality freezing of tissue samples in thickness ranges of up to 0.2 mm without requiring freeze protective additives.
• Array of specimen carriers available, i.e. for suspensions (capillary tubes), tissue extracted by fine needle biopsy and carriers for monolayer cell cultures.
• In-situ real time measurement of temperature and pressure.
• Fast 90 second process cycle allows expeditious application.
• Automatic, microprocessor controlled operation for routine work.
• Compact, sturdy unit with soundproof and vibration-free housing.
• Simple and safe operation due to quick-locking action of specimen holder and clearly arranged operational controls.
• Processing datas are recorded on digital display, such as the actual temperature, time and pressure, thus allowing the user to exactly evaluate current operational status (sample quality control).
• Officially approved materials and high pressure components, as well as twice controlled sample holders with quick-lock action ensure highest level of safety for user.
• Simple maintenance with removable cover plates and rack system for control units.
• Extensive accessory program.
• Large number of publications documenting high performance of HPM 010.

Applications

The High Pressure Freezing Method

Excellent freezing of the specimens intended for examination in the electron microscope is one of the most important prerequisites for achieving reproducible results from the various subsequent cryopreparation methods.

The freezing method should produce microcrystalline or amorphous ice from the specimen water. To achieve this, the specimens must be frozen as quickly as possible at a freezing rate no lower than 10’ 000°C/s.

The conventional freezing methods in use are plunge freezing, jet spray and cold block (slamming) cryo fixation. However, due to the poor heat conductance of water, these methods can only satisfactorily freeze specimens measuring up to between 10 and 20µm.

Thicker specimens (such as tissue samples) could only be frozen in the past, if a cryoprotectant was added to lower the freezing point of the water in the specimen. The disadvantage of chemical cryoprotectants is that they often affect certain cell structures, causing different types of undesirable artefacts.

By using the high pressure freezing method developed by Prof. Moor (ETH Zurich) in conjunction with BAL-TEC AG these artefacts can be eliminated. The high pressure method is based on an entirely physical phenomenon that lowers the freezing point of water. This method functions as follows: At 2’ 100 bar the melting point of water drops from 0°C to -22°C. Under normal atmospheric conditions homogeneous nucleation (supercooling) begins at -40°C. Under high pressure this nucleation doesn’t begin until the water has reached -90°C (see H20 Phase Diagram).

At 2’ 100 bar water is 1’ 500 times more viscous than at atmospheric pressure, which drastically reduces the nucleation and thus the crystal growth rate. This means that the extremely high freezing rate (min. 10’ 000°C/s required for satisfactory freezing by the methods previously mentioned is not necessary with the high pressure method. The high pressure method allows specimens up to 0.2mm thickness with a total volume of approx. 1mm 3 to be vitrified or up to 0.5mm thickness to be adequate frozen at a low freezing rate of 200°C/s without requiring the addition of cryoprotectants.