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Critical Point Drying Preparation Techniques and Advantages

Here you can find information and downloads to explain the techniques and advantages of critical point drying. Details on this page cover:

A summary of the critical point drying method

Critical point drying is an established method of dehydrating biological tissue prior to examination in the Scanning Electron Microscope. The technique was first introduced commercially for SEM specimen preparation by Polaron Ltd in 1971. The original design concepts, which included a horizontal chamber, are still embodied in the design of the E3000 and E3100 CPD models.

In recent years we have introduced two further models: the K850, which features built-in chamber cooling and heating, and the K850WM, which is designed for drying a 100mm/4” silicon wafer.

All three models have found general acceptance in many laboratories throughout the world. Together, these critical point dryers offer the user a choice most suited to the particular specimen preparation requirements.

The phase diagram shows the pressure to temperature ranges where solid, liquid and vapour exist. The boundaries between the phases meet at a point on the graph called the triple point. Along the boundary between the liquid and vapour phases it is possible to choose a particular temperature and corresponding pressure, where liquid and vapour can co-exist and hence have the same density. This is the critical temperature and pressure.

Critical point drying relies on this physical principle. The water in biological tissue is replaced with a suitable inert fluid whose critical temperature for a realisable pressure is just above ambient. The choice of fluids is severely limited and CO2 is universally used today, despite early work with Freon 13 and nitrous oxide.

With CO2 a critical point of approximately 35°C can be achieved at a pressure of around 1,200psi. Therefore if the water is replaced with liquid CO2 and the temperature then raised to above the critical temperature, the liquid CO2 changes to vapour without change of density and therefore without surface tension effects which distort morphology and ultra structure.

Since liquid CO2 is not sufficiently miscible with water, it is necessary to use an intermediate fluid which is miscible with both water and liquid CO2. In practice intermediate fluids commonly used are methanol, ethanol, amyl acetate and acetone.

CO2 grades required for critical point drying

Generally speaking, the grade we recommend is ‘normal’ grade - that is the one most commonly offered by industrial gas suppliers.

In most parts of the world, ‘normal’ grade of CO2, from the suppliers, is specified as ‘N4.5’ or 99.995% minimum purity with a maximum level of 50ppm of impurities.

There are, however, two other grades available by special order from most gas suppliers. One is ‘N4.0’ or 99.99% (less pure than ‘normal’ grade). The other is ‘N5.5’ or 99.9995% minimum purity. The N5.5 purity is easier to find in those parts of the world where there is high level of activity in electronics, since these customers often demand gases with higher purities.

We are not aware of anyone who has ever reported either superior results using N5.5 purity vs N4.5, or inferior results using N4.5 vs N5.5. However, we do want our customers to have the benefit of such detail about liquid carbon CO2 procurement in the event they should ever find reason to believe that their particular specimens might benefit from the higher purity product.

Remember, the requirement is for LIQUID carbon dioxide and NOT gaseous carbon dioxide. For this reason a cylinder with an internal ‘siphon’ must be specified. A siphon cylinder is normally denoted by a white stripe painted along its length. No pressure regular is needed.

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