The Paraffin bath



This step is usually called infiltration, but when the transfer from the clearing fluid to paraffin is made gradually, as has just been indicated, the process of infiltration is already begun. It is now necessary to get rid of the xylol or other clearing agent. This is accomplished, to a considerable extent, by pouring off the mixture of xylol and paraffin and replacing it with pure melted paraffin. Pour off the pure paraffin immediately. This is important. You will notice that often, when the pure paraffin is poured on, a froth or scum will appear on the surface. Much of the xylol will be in this scum, and, if allowed to remain, it would diffuse into the mass and greatly prolong the time needed for infiltration. So, pour it off and add more pure paraffin, for some xylol remains in the tissues and must be removed. Do not put the shell into the bath, but use a flat dish of some sort. The main object is to have a fairly large surface exposed, so that the remaining xylol may evaporate as rapidly as possible. Change the paraffin 2 or 3 times. Soft paraffin (about 45 C.) may be used at first, but the second should be the paraffin of the grade in which the material is to be imbedded. If there are two baths, one should be kept at 46 C. and the other at 52 C., if the material is to be imbedded in 52 C. paraffin. While using the soft paraffin, keep the material in the 46 C. bath; for the harder paraffin, use the 52 C. bath. If there is only one bath, there is no object in using the 45 C. paraffin. Do not throw away the paraffin which you pour off, but put it in a waste jar or beaker, or, still better, in a small tin lard pail, in which you have made a lip to facilitate pouring. This can be placed in the bath, or, in winter, on the radiator, and the xylol will gradually evaporate. After long heating, the paraffin not only becomes as good as new, but even better, since it becomes more homogeneous and tenacious. If it contains dust or debris of any kind, it may be filtered with a hot filter. The time required varies with the character of the material and the thoroughness of the dehydrating and clearing. If this schedule has been followed up to this point, the time will be much shorter than most investigators now deem necessary. Fern prothallia infiltrate perfectly in 15 to 20 minutes; onion root-tips in 20 to 30 minutes; ovaries of Lilium at the fertilization stage, 30 minutes to 1 hour; 5 or 6 mm. cubes of endosperm of cycads, containing archegonia, 2 to 21 hours; median longitudinal sections, 4 or 5 mm. thick, through ovulate cones of Pinus Banksiana may require 6 or 8 hours; if serial sections through the entire cone are wanted, Miss Aase found that the time must be prolonged to 24 or even 48 hours. When one is dealing with many lots of the same kind of material, as in research work, the time required for infiltration is easily determined. As a rule, minimize heat. It is, probably, never necessary to use paraffin with a melting-point higher than 52 C. With Land's cooling device sections 1( in thickness can be cut from 52 C. paraffin.





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