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ICC (immunocytochemistry) Applications
of Microwave Processing

The use of the microwave to enhance or accelerate immunocytochemistry protocols (i.e. direct, indirect or kits from Vector Labs or DAKO as examples) have been published (see Kok and Boon, 1992, Microwave Cookbook for Microscopists, 3rd ed. Coulomb Press, Leyden, ch. 17). However, there is little continuity between the various protocols leading to confusion as to the role of the microwave in the process.

A. The following is a direct quotation from the reference listed above relating to the use of microwave irradiation for ICC:

"Notwithstanding these positive reports, we certainly do not consider the application of microwave exposure to be simple. The linking of antibody to antigen is often a very delicate process and many complexes prove to be unstable when the temperature becomes too high and when the microwave exposure too intense. Therefore, to perform immunoreactions in the microwave oven, certain precautions must be taken first to keep the system in hand and some steps (with instable complexes) must be performed outside the microwave oven."

B. It is evident, from the above quote, that control of the microwave environment is central to its use during ICC. Variable wattage controls the intensity of the microwave in the chamber as well as the heating rate. The temperature probe controls temperature maximums reached. The water recirculator creates a uniform environment for all slides being stained. By continuous cooling of the water under the slides, the microwave stays on almost continuously during staining and accurate temperature control of all slides is maintained.

C. Staining is done by means of a slide rack immersed in a water bath which is maintained at 40°C by the water recirculator. The temperature probe insures that the maximum temperature, where the slides are, does not exceed 40°C. The microwave is programmed to run for 2 minutes on at 100% power, 2 minutes off (0% power) and 2 minutes on at 100% power for each ICC step. The rinse is done outside the microwave. If the ICC protocol is an indirect procedure, the process is repeated. Development of the chromagen can be done in the microwave under the same conditions, except it is run as a continuous process (typically 1-2 minutes at 100% power).

D. Wattage is set at 550 or 650 watts.

E. Microwave-assisted ICC protocols typically produce results having low background and equal or better positive staining, when compared to routine or automated ICC protocols.

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