003591 53/87/620l-0059$02.00/0 STAIN TECHNOL~CY Copyright Q 1987 by The Williams & Wilkins Co.

Vol. 62. No. 1 Prinud in U. S. A.

NOTES ON TECHNIC

ALUMINUM FOIL MOLDS FOR CRYOSTAT BLOCKS

GIAN-LUCA FERRI, CARLA PAPADIA, DOMENICO COCCHIA AND JULIA M. POLAK, Department of Human Anatomy, Tor Vergata University, 00173 Rome, Italy, and Department of Histochemistry, Royal Postgraduate

Medical School, Hammersmith Hospital, London W12 United Kingdom

Inexpensive pyramidal templates may be prepared in a range of sizes from segments of plastic or wooden bars (Fig. 1). Pieces of double-layered aluminum cooking foil are shaped by hand around templates of appropriate size and their surfaces are smoothed with fingernails. In this way, "negative" molds of known cross-section and variable depth can be obtained (Fig. 1). Tissue samples are oriented on the bottom of the molds in a few drops of Ames O.C.T. Compound and just covered with the same embedding medium. Narrow strips of light-weight cardboard with identification data are inserted into the blocks close to a lateral surface. The blocks are solidified on the cold stage of a cryostat (a drop of O.C.T. will improve thermal contact and help keep them upright), or with a liquid gas, such as nitrogen, or Freon 22 (in the latter case the molds are immersed as far as the upper level of the O.C.T., but are not submerged, so that the Freon does not bubble into the liquid O.C.T.). After freezing, the foil is removed and the blocks, handled with cold forceps, are mounted on cryostat chucks with O.C.T.

The above method enabled us to prepare quickly high quality blocks of virtually every tissue tested. By using molds that closely fit the tissue samples, freezing time can be kept almost as short as that for tissue alone, thus minimizing tissue damage due to ice crystal formation (Pearse 1980). Due to the high thermal conductivity of aluminum, the present method was preferred to similar ones using materials of low thermal conductivity such as polyethylene (Fasano et al. 1974), polystyrene (Shannon 1973) or paper (Gabe 1976) for mold construction. The specimen holder of our cryomicrotome (Leitz Model 1720) was oriented so that the flat front surface of blocks was aligned with the knife edge. In this way, after a few turns of the cryostat handle, the whole face of the block and the included specimen was cut.

For most tissues, setting both chamber and specimen temperatures between -25 and -30 C ( i e . , 5-10 C lower than we normally use when cutting at I0 pm) was of significant help in producing ribbons of sections of first quality down to 3 pm thick. Using two small brushes kept cold in the crymtat chamber, single adjacent sections could be detached and mounted on the same or separate slides whenever required for comparative study. Finally, the exces O.C.T. used for mounting was safely trimmed by cutting around the block base, thus leaving blocks of virtually the starting size ready for storage in limited freezer space.

59

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60 STAIN TECHNOLOGY

FIG. 1. Plastic templates (tip size, in millimeters, k indicated around the hase), aluminum foil molds and frozen blocks. One of the embedded tissue blocks is mounted on a chuck fitted on a freezing wage of our Lcitz cryomicrotome. Note the narrow base of the pyramidal templates, their lateral faces being almost parallel, and the lateral location of the cardboard label, which facilitates mounting.

REFERENCES Fasano. A. V.. &rendsen. P. B. and Labriob. A. 1974. T h e use of BEEM capsules in the

Cabe. M. 1976. HU&&gid Techniques. Blackith. R. E. and Kovoor, A. (trs.). Springer, Berlin, Parse. A. C. E. 1980. His&chistr), Threticul and Applied, 4th ed. Vol. 1 (Preparative and

Shannon, W. A. 1973. Tissue molds for cryostat embedding. Stain Technol. 48: 266-268.

preparation of fresh frozen cryatat sections. Stain Technol. 49: 55-55.

Optiil Technology). Churchill Livingtone, Edinburgh.

A SIMPLE METHOD FOR RAPID DETECTION OF CHROMOSOMES OR NUCLEI IN YOLK-RICH AVIANOOCYTES OR EGGS

MARC CALLEBAUT AND ROBERT-JAN SIJENS, Laboratory ofAnatomy and Embryology, State Univcrsiry of Antwerp, B-2020 Antwerpen, Belgium

As early as 19 14 Van Durme described how difficult it is to detect the small avian chromosomes among the enormous mass of yolk granules at the end of oogenesis. During the ensuing fertilization and early cleavage period, he could hardly follow the rapidly changing morphology of the nuclei and chromo- somes, since the classical staining methods also stained the intervitelline material.

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