2. Smears. A clean fresh cut is made into the organ or tissue, and a clean slide or cover-glass is drawn across the surface. Without permitting the smear to dry a drop of salt-solution or any desired reagent is put upon it, and it is then examined. This method is especially applicable to the study of the cells of the spleen, bone-marrow, lymphnodes, etc. Permanent balsam-mounts may be made of such smears by fixing with heat or alcohol and ether, staining, drying and mounting.
3. Scraping. A fresh cut is made into the organ or tissue, and the excess of blood absorbed by a pad of absorbent paper devoid of lint. A clean scalpel held at an angle of 45° is then drawn with some force back and forth over the cut-surface until its blade collects a sufficient amount of “tissue-juice” made up of the cells of the tissue. This is then put upon a slide, and covered with salt-solution or any desired reagent, and is then examined. This method is used especially for cellular infiltrations, soft tumors, and parenchymatous organs (spleen, lymphnodes, bone-marrow, liver, etc.), and for the inner wall of cysts (echinococcus-cysts, cysts lined with ciliated epithelium).
4. Crushing. A small bit of the tissue is cut out with the scissors or scalpel, and placed upon the slide. A cover-glass is then placed upon it and pressed down so firmly that the bit of tissue is spread out in a thin film or layer beneath the cover-glass. Reagents are introduced beneath the cover-glass, as desired. (See below.) This method is used in the examination of the lung, kidneys and brain for fatty embolism, and of the brain and spinal-cord for “fat-granule” cells, pigment, calcified ganglion-cells, Negri bodies, etc. It is also frequently used in the bacteriologic examination of tissues (crushing of tubercles, etc.).
5. Teasing. A small bit of the tissue is cut out and placed upon the slide or in a staining-dish and covered with physiologic salt-solution. It is then teased with fine needles until divided into its ultimate elements. Hard tumors (mature connective-tissue tumors, fibrosarcomata, etc.), muscle (examination for trichina), nerve-trunks, etc., are best examined in this way.
6. Maceration. In the case of some tissues the ultimate histologic elements are so firmly held together that they cannot be separated without the aid of a macerating- or dissociation-fluid. (See below.) Digestion is also used for the same end. The tissue should be as fresh as possible and cut into small bits, which are placed in the maceration-fluid in watch-glasses or staining-dishes for twenty-four hours or longer. The macerated bits are then teased until the finest elements are separated. In the case of very minute elements the teasing may be carried on under a hand-lens or the stereoscopic binocular microscope. During the process of maceration all parts of the macerating tissue must be kept covered with the macerating-fluid, or the uncovered portions will become hardened.
7. Section-cutting. Sections of fresh tissue may be made with the curved shears, simple razor, double-bladed razor and the freezing-microtome.
a. Curved Shears. The tissue is put upon a stretch, and from the surface a thin, flat section is cut out with the scissors. With care a fairly thin section may be obtained in this way. It may be examined by pressing it upon a slide beneath a cover-glass, or it may be treated as a frozen section.
b. Simple Razor. As a part of their required laboratory training medical students should learn how to make working sections with a simple razor. Such a technical knowledge is sure to be of practical use at some time or other. With a little practice sections sufficiently thin for ordinary diagnostic work can be cut. If the piece of tissue is large it may be held in the hand; but when small or soft it may be placed in a matrix of hardened liver, pith, potato, apple, firm lard or butter, paraffin, etc., and cut at the same time with the latter. Both blade and tissue should be wet with physiologic salt-solution. The blade, which must be very sharp, should be drawn through the tissue by a shoulder movement, with the wrist-joint fixed. As the sections are cut they are floated off of the razor-blade into physiologic salt-solution, and thence treated as desired.
c. Double-bladed Knife. This consists of two parallel blades arranged so that the space between them can be changed by means of screws. For firm material the blades should be close together; for soft material farther apart. The blades should be dipped into physiologic salt-solution to fill up the space between the blades. The instrument is then drawn through the tissue, and the section between the blades floated out in salt-solution. Some workers use this double-bladed knife to make a perpendicular cut into the tissue, then turning it to either side to cut the lower edge of the section and removing the blades with the section between them. Both the single- and the double-bladed razors require practice for successful section-cutting. Further, it must be borne in mind that sections of fresh tissue obtained by these methods are unsuited for complicated staining methods and can be used only for the simplest staining processes. Nevertheless, in a fairly large proportion of pathologic conditions it is possible to secure a diagnosis by these methods.
d. Freezing Microtome. Much more satisfactory sections can be obtained by freezing the fresh tissue and cutting it upon the freezing microtome. Various types of these instruments are in the market. The freezing is accomplished by the use of ether, ethyl-chloride or fluid carbonic acid gas. The ether and ethyl-chloride freezing-microtomes consist essentially of a metal plate or hollow box on which the tissue rests and against the under-surface of which a spray of ether or ethyl-chloride is forced by means of a rubber bulb connected with a supply of the freezing agent in a bottle. Such freezing attachments can be attached to any microtome, but special instruments as Jung’s “student’s freezing microtome,” Cathcart’s, Bausch and Lomb’s or Becker’s ether-freezing microtome can be recommended for this purpose. (See Fig. [52].) The tissue to be frozen must be of small size and not more than 3-4 mm. thick. It is placed upon the freezing plate in a drop of water, white of egg or thick gum-arabic, and pressed firmly against the plate. The spray must not be too constant or strong, but should be given with regular pauses of about a second to allow the ether to evaporate. The tissue must be firmly frozen, but not so hard that it crumbles. It is usually difficult to freeze the upper part of the tissue hard enough to give good sections, and this half-frozen tissue must be trimmed off with the knife until good sections are secured. As the sections are cut they are removed from the knife-blade with the finger and put into physiologic salt-solution that has been recently boiled to drive off the air, so that, in thawing, there may be no formation of air-bubbles in the tissue to cause artefacts. If put into strong alcohol diffusion-currents may damage the tissue; a succession of graded solutions should, therefore, be used if the sections are to be fixed and hardened. The further treatment of frozen sections is given below. The relative slowness of the method of freezing, its greater cost, the necessity of the frequent replacement of the rubber bulbs and tubing, and the greater amount of trouble required by the use of ether or ethyl-chloride are disadvantages that can be avoided by the use of the carbonic acid freezing microtome. Where much work is done by the freezing method the use of the latter is advised.