LESSON II.

THE BLOOD.

UNDER the microscope blood is seen to consist of a clear, transparent fluid, the plasma or liquor sanguinis, in which are suspended the blood-corpuscles. The blood-corpuscles are of two kinds, the red or coloured, and the white, pale, or colourless. Besides these, there fall to be examined the blood-plates, or, as they are also called, blood-tablets or platelets.

(A.) Coloured Corpuscles of Amphibians (Newt or Frog).-After destroying the brain of a newt or frog, a drop of blood may be obtained from the cut end of the tail of the former, or from the cut surface after amputation of the foot of the latter.

1. Red Corpuscles (H).-Place a small drop of blood in the centre of a perfectly clean slide and cover it at once with a cover

glass. Examine it with a high power. To avoid the pressure of

the cover-glass, a short length of a hair may be placed in the blood droplet before the cover-glass is applied.

(a.) Observe the coloured and colourless corpuscles, the former much more numerous than the latter (fig. 61).

(b.) Study the red corpuscles. Observe that they are very numerous, elliptical in outline when seen on the flat, slightly yellowish in colour; their border or contour is even and well defined. Select one seen on edge, and note that it is a thin ellipse, pointed at the ends, becoming gradually thicker in the centre, so that it is a bi-convex elliptical disc. Sometimes one corpuscle can be seen overlying a subjacent one, in which case the outline of the latter can be distinctly seen through the former, indicating that the corpuscles are transparent (fig. 61). Notice within each corpuscle a lighter oval, central area, indicating the existence of an elliptical, colourless,

n

elongated, granular-looking included body--the nucleus. The long axis of the colourless nucleus coincides with the long axis of the corpuscle. At first, the nucleus may not be very distinct, but after a time it becomes distinctly visible, and it can be readily made so by the action of certain reagents, especially weak acids. Small vacuoles frequently appear in the body of the corpuscle, more especially in frogs that have been kept some time. In others there

may be seen a faint radiate striping of the corpuscles as if there were fine folds in it due to partial drying.

The yellow colour is due to the presence of hæmoglobin, which is enclosed within the meshes of a colourless stroma or delicate framework.

If the blood be taken from a frog which has been kept throughout the winter, vacuoles may be

FIG. 61.-Blood of Frog. a. Red corpuscle seen on the flat; b. In Profile; c. Threequarter face; some of the red corpuscles show the presence of vacuoles (v); n. Colourless corpuscle at rest; m. One with amoeboid processes; p. Fusiform cell, probably from the vascular wall.

seen in the hæmoglobin of the red corpuscles (fig. 61, v). They are rapidly produced in frogs after the injection of ammonium chloride. Sketch two or three corpuscles, both red and white; the red corpuscles both on the flat and on edge, and two

overlapping each other.

-I CC.

2

2. Acetic Acid. To the edge of the cover-glass of the same preparation apply a drop of dilute acetic acid (1 per cent.glacial acetic acid to 99 cc. of normal saline solution). The acid runs in under the coverglass, but if it does not, apply a small triangular piece of blotting-paper to the opposite edge of the cover-glass. Let one of the angles of the blotting-paper touch the fluid under the cover-glass, and it will suck up some of the blood, and thus cause the acetic acid to run in at the opposite side. This is known as the process of irrigation. Move the slide, and find a part of the object which has been acted on by the dilute acid. (a.) Observe that the red corpuscles, or at least most of them, become spherical and decolorised, the nucleus becomes very distinct

X 1000.

and granular, while the outline of the corpuscle may become very indistinct (fig. 63). The nucleus appears somewhat shrunken or shrivelled. If the corpuscles are roughly treated, the nucleus may be seen placed excentrically, or even extruded from the corpuscle. The acid acts on the hæmoglobin, forming a new compound, which diffuses out of the corpuscles and stains the surrounding plasma a faint yellow. The nucleus in some of the corpuscles may absorb some of the yellow pigment, and become stained thereby, especially if a strong solution of acid has been used. The action on the colourless corpuscles is referred to at p. 112.

3. Dilute Hydrochloric Acid, 1 per cent. (H).

(a.) To a fresh drop of blood add, as before, a drop or two of dilute hydrochloric acid. Watch diligently one or two of the red corpuscles. They gradually enlarge, become spherical, and may all of a sudden burst and discharge their contents, the nucleus coming clearly into view during the process. After the rupture, the residue of the stroma of the corpuscles may be seen in the field. In other cases the corpuscles become clear, globular, and transparent, with

FIG. 63.-Frog's Red
Blood Corpuscle
acted on by Dilute
Acetic Acid, x 300.

FIG. 64. Action of

Water on an Am-
phibian Coloured
Blood-Corpuscle.

FIG. 65. Action of
Syrup on Frog's
Red Blood - Cor-
puscle, × 300.

here and there fine shreds stretching between the nucleus and the surface of the spherical corpuscle.

4. Water (H). To a fresh preparation of blood apply a drop of water to the edge of the cover-glass, and notice its effects upon the corpuscles.

(a.) The water rapidly diffuses into the corpuscles and renders them spherical, while at the same time it decolorises them, the hæmoglobin diffusing outwards into the plasma, and staining it. slightly yellow. The nucleus also becomes spherical. Thus the outline of the corpuscles becomes very faint in the field of the microscope, the corpuscles themselves now almost consisting of a nucleated stroma (fig. 64).

5. Strong Syrup (4).—Place a small drop of blood on a slide and near it a drop of syrup; mix the two with a needle, and apply a cover-glass.

(a.) Observe that some of the red corpuscles are rapidly shrivelled and puckered, especially when seen on edge, owing to fluid passing out of them by exosmosis (fig. 65). Some of them

may present here and there a reddish tinge. are not affected equally or at the same time. produced by strong saline solutions.

All the corpuscles The same effects are

6. Tannic Acid (H).-On a slide mix a drop of blood with a drop of tannic acid, using a relatively large drop of the acid fluid; wait for about a minute, and then apply a cover-glass.

(a.) Observe that some of the corpuscles become globular, while the hæmoglobin passes out of the corpuscles at one or more spots, and appears on the surface in the form of one or more small granular buds (fig. 66, c). Sometimes the bud or buds are small. At others the bud may be as large as the remainder of the corpuscle, which has become smaller and partly decolorised. In other cases

it may be collected around the nucleus (Roberts). The tannic acid causes a separation of the hæmoglobin from the stroma.

The solution of tannic acid is made by dissolving 2 grains of tannic acid in 1 oz. of boiling water, and allowing it to cool.

FIG. 66.-Action of Tannic Acid

on the Red Blood-Corpuscles
of Man (a, b) and Frog (c).

FIG. 67.-Action of Boracic Acid on a Frog's
Red Blood-Corpuscle.

7. Boracic Acid.-Mix a drop of newt's or salamander's blood with a 2 per cent. solution of boracic acid, which takes a short time to act on the corpuscles and produce its effects.

(a.) Observe that the hæmoglobin is collected around the nucleus, so that the presence of the latter is obscured, but in many of the corpuscles fine threads of hæmoglobin remain attached to the circumference of the corpuscle, so that the retracted hæmoglobin may have a stellate form, while the rest of the corpuscle is colourless. To the latter Brücke gave the name Oikoid, to the former Zooid (fig. 67).

If these corpuscles are acted on for twenty-four hours with 1 per cent. osmic acid, they are "fixed," and may be mounted permanently in glycerine-jelly.

8. Magenta. Mix a drop of blood and a drop of the special magenta fluid (p. 74) on a slide, cover, and examine.

(a.) Observe that the nuclei of the corpuscles, both red and white, are stained of a brilliant red, although the surrounding part of the corpuscle is not so stained, unless the magenta be in great excess. All the corpuscles are not stained equally brightly. On the edge of some of the corpuscles at one or more points will be found small coloured spots or thickenings.

9. Osmic Acid and Picrocarmine (H).-This preparation is best made by mixing a few drops of blood with an equal volume of I per cent. osmic acid in a small tightly-corked tube, or by exposing a thin film of blood to the vapour of a 2 per cent. solution of osmic acid. After two to four hours pour off the supernatant fluid and cover the residue of corpuscles with picrocarmine. After twentyfour hours the picrocarmine can be poured off, and a little of the deposit placed on a slide and mixed with glycerine-jelly dissolved by heat, and covered (p. 85).

(a.) Observe that the nuclei of the corpuscles are bright-red, and the perinuclear part of the corpuscles yellow. Within the granularlooking nucleus, with a good lens, may be seen a network of fibrils. This preparation is permanent, and must be sealed up or “ringed" after the manner described at p. 88.

10. Blood of Bird (H).—Mount a drop of fresh blood.

(a.) Observe the red corpuscles, which are elliptical, biconvex, nucleated bodies, but smaller in size and more pointed than amphibian corpuscles (fig. 68).

These corpuscles behave towards reagents as those of amphibians. 11. Blood of Fish (H).—Mount a drop of fresh blood, which is readily obtained from a gold-fish or salmon, but the blood coagulates rapidly.

(a.) Observe the red corpuscles, which are elliptical, biconvex, and nucleated, but the ends are not so pointed as in the bird, while,

like the bird's corpuscles, they are smaller than those of amphibians (fig. 68, F). The blood must

be quite fresh.

In both cases a few colourless corpuscles may be noticed.

(B.) The White Corpuscles of the Frog or Newt.

12. White Corpuscles. In a fresh preparation of blood (taking care to place a hair under the cover-glass) search for the colourless corpuscles, of which there are several varieties. They are much less numerous than the red.

(a.) Observe that there is: (i.) The finely granular form, consisting of a nucleated mass of protoplasm larger than a red corpuscle (fig. 61, m). (i.) The coarsely granular variety may be found. In it the granules are large and refractive, and often lying at one side. of the corpuscles. (iii.) A third variety, much smaller than the others, may be found (fig. 61, k).