Serum-alb.u.min gives all the typical colour and precipitation reactions of the alb.u.mins. If plasma be weakly acidified with sulphuric acid, then treated with crystals of ammonium sulphate until a slight precipitate forms, filtered and the filtrate allowed to evaporate very slowly, typical crystals of serum-alb.u.min may form. According to many it is a uniform and specific substance, but others hold the view that it consists of at least three distinct substances, as shown by the fact that if a solution be gradually heated coagulation will occur at three different temperatures, viz. at 73, 77 and 84 C. On the other hand the close agreement between different a.n.a.lyses of even the amorphous preparations points to there being but one serum-alb.u.min.

When blood clots two new proteins make their appearance in the fluid part of the blood, or serum, as it is now called. The first of these is fibrin ferment (for its origin see section on _Clotting_ below). The other, fibrinoglobulin, possesses all the typical characteristics of the globulins and coagulates at 64 C.

_Carbohydrates._--Three several carbohydrates are described as occurring in plasma, viz. glycogen, animal gum and dextrose. If glycogen is present in solution in the plasma it is there in very small quant.i.ties only, and has probably arisen from the destruction of the white blood corpuscles, since some leucocytes undoubtedly contain glycogen. A small amount of carbohydrate having the formula for starch and yielding a reducing sugar on hydrolysis with acid has also been described. The constant carbohydrate const.i.tuent of plasma, however, is dextrose. This is present to the approximate amount of 0.15% in arterial blood. The amount may be much greater in the blood of the portal vein during carbohydrate absorption, and according to some observers there is less in venous than in arterial blood, but the difference is small and falls within the error of observation. The statement that when no absorption is taking place the blood of the hepatic vein is richer in dextrose than that of the portal vein (Bernard) is denied by Pavy.

_Fats._--Plasma or serum is as a rule quite clear, but after a meal rich in fats it may become quite milky owing to the presence of neutral fats in a very fine state of subdivision. This suspended fat rapidly disappears from the blood after fat absorption has ceased. To some extent it varies in composition with that of the fat absorbed, but usually consists of the glycerides of the common fatty acids--palmitic, stearic and oleic. In addition, there is a small amount of fatty acid in solution in the plasma. As to the form in which this occurs there is some uncertainty. It is possibly present as a soap or even as a neutral fat, since a little can be dissolved in plasma, the solvent substance being probably protein or cholesterin. Fatty acids also appear to be present to some extent combined with cholesterin forming cholesterin esters (about 0.06%).

_Other Organic Compounds._--In addition to the substances above described, belonging to the three main cla.s.ses of food stuffs, there are still other organic bodies present in plasma in small amounts, which for convenience we may cla.s.sify as non-nitrogenous and nitrogenous. Among the former may be mentioned lactic acid, glycerin, a lipochrome, and probably many other substances of a similar type whose separation has not yet been effected.

The non-protein nitrogenous const.i.tuents consist of the following: ammonia as carbonate or carbamate (0.2 to 0.6%), urea (0.02 to 0.05%), creatine, creatinine, uric acid, xanthine, hypoxanthine and occasionally hippuric acid. Three ferments are also described as being present: (1) a glycolytic ferment exerting an action upon dextrose; (2) a lipase or fat-splitting ferment; and (3) a diastase capable of converting starch into sugar.

_Salts._--The saline const.i.tuents of plasma comprise chlorides, phosphates, carbonates and possibly sulphates, of sodium, pota.s.sium, calcium and magnesium. The most abundant metal is sodium and the most abundant acid is hydrochloric. These two are present in sufficient amount to form about 0.65% of sodium chloride. The phosphate is present to about 0.02%. Sulphuric acid is always present if the blood has been calcined for the purposes of the a.n.a.lysis, and may then be present to about 0.013%. This is, however, probably produced during the destruction of the protein, since it has been shown that no sulphate can be removed from normal plasma by dialysis. The amount of pota.s.sium present (0.03%) is less than one-tenth of that of the sodium, and the quant.i.ties of calcium and magnesium are even less.

_Formed Elements._--When viewed under the microscope the main number of these are seen to be small yellow bodies of very uniform size, size and shape varying, however, in different animals. When observed in bulk they have a red colour, their presence in fact giving the typical colour to blood. These are the _red blood corpuscles_ or _erythrocytes_ (Gr.

[Greek: erythros], red). Mingled with them in the blood are a smaller number of corpuscles which possess no colour and have therefore been called _white blood corpuscles_ or _leucocytes_ (Gr. [Greek: leukos], white). Lastly, there are present a large number of small lens-shaped structures, less in number than the red corpuscles, and much more difficult to distinguish. These are known as _blood platelets_.

_Red Corpuscles._--These are present in very large numbers and, under normal conditions, all possess exactly the same appearance. With rare exceptions their shape is that of a biconcave disk with bevelled edges, the size varying somewhat in different animals, as is seen in the following table which gives their diameters:--

Man 0.0075 mm.

Dog 0.0073 mm.

Rabbit 0.0069 mm.

Cat 0.0065 mm.

Goat 0.0041 mm.

The coloured corpuscles of amphibia as well as of nearly all vertebrates below mammals are biconvex and elliptical. The following are the dimensions of some of the more common:--

Pigeon 0.0147 mm. long by 0.0065 mm. wide.

Frog 0.0223 " " 0.0157 " "

Newt 0.0293 " " 0.0195 " "

Proteus 0.0580 " " 0.0350 " "

Amphiuma 0.0770 " " 0.0460 " "

Their number also varies as follows:--

Man 4,000,000 to 5,000,000 per cub. mm.

Goat 9,000,000 to 10,000,000 " "

Sheep 13,000,000 to 14,000,000 " "

Birds 1,000,000 to 4,000,000 " "

Fish 250,000 to 2,000,000 " "

Frog 500,000 per cub. mm.

Proteus 36,000 " "

In mammals they are apparently h.o.m.ogeneous in structure, have no nucleus, but possess a thin envelope. Their specific gravity is distinctly higher than that of the plasma (1.088), so that if clotting has been prevented, blood on standing yields a large deposit which may form as much as half the total volume of the blood.

_Chemical Composition._--On destruction the red corpuscles yield two chief proteins, haemoglobin and a nucleo-protein, and a number of other substances similar to those usually obtained on the break-down of any cellular tissue, such for instance as lecithin, cholesterin and inorganic salts. The most important protein is the haemoglobin. To it the corpuscle owes its distinctive property of acting as an oxygen carrier, for it possesses the power of combining chemically with oxygen and of yielding up that same oxygen whenever there is a decrease in the concentration of the oxygen in the solvent. Thus in a given solution of haemoglobin the amount of it which is combined with oxygen depends absolutely on the oxygen concentration. The greatest dissociation of oxyhaemoglobin occurs as the oxygen tension falls from about 40 to 20 mm. of mercury. That the oxygen forms a definite compound with the haemoglobin is proved by the fact that haemoglobin thoroughly saturated with oxygen (oxyhaemoglobin) has a definite absorption spectrum showing two bands between the D and E lines, whilst haemoglobin from which the oxygen has been completely removed only gives one band between those lines. In a.s.sociation with this, oxyhaemoglobin has a typical bright red colour, whereas haemoglobin is dark purple. A further striking characteristic of haemoglobin is that it contains iron in its molecule.

The amount present, though small bears a perfectly definite quant.i.tative relation to the amount of oxygen with which the haemoglobin is capable of combining (two atoms of oxygen to one of iron). One gram of haemoglobin crystals can combine with 1.34 cc. of oxygen. On destruction with an acid or alkali, haemoglobin yields a pigment portion, haematin, and a protein portion, globin, the latter belonging to the group of the histones (Gr. [Greek: istos], web, tissue). In this cleavage the iron is found in the pigment. By the use of a strong acid, it may be made to yield iron-free pigment, the remainder of the molecule being much further decomposed.

_Destruction and Formation._--In the performance of their work the corpuscles gradually deteriorate. They are then destroyed, chiefly in the liver, but whether the whole of this process is effected by the liver alone is not decided. It is proved, however, that the destruction of the haemoglobin is entirely effected there. It was for a long time considered to be one of the functions of the spleen to examine the red corpuscles and to destroy or in some way to mark those no longer fitted for the performance of their work. It is proved that the destruction of the haemoglobin is entirely effected in the liver, since both the main cleavage products may be traced to this organ, which discharges the pigmentary portion as the bile pigment, but retains the iron-protein moiety at any rate for a time. The amount of bile pigment eliminated during the day indicates that the destruction must be considerable, and since the number of corpuscles does not vary there must be an equivalent formation of new ones. This takes place in the red bone-marrow, where special cells are provided for their continuous production. In embryonic life their formation is effected in another way. Certain mesodermic cells, resembling those of the connective tissue, collect ma.s.ses of haemoglobin, and from these elaborate red blood corpuscles which thus come to lie in the fluid part of the cell. By a ca.n.a.lization of the branches of these cells which unite with branches of other cells the precursors of the blood capillaries are formed.

_White Blood Corpuscles._--These const.i.tute the second important group of formed elements in the blood, and number about 12,000 to 20,000 per cubic mm. They are typical wandering cells carried to all parts of the body by the blood stream, but often leave that stream and gain the tissue s.p.a.ces by pa.s.sing through the capillary wall. They exist in many varieties and were first cla.s.sified according as, under the microscope, they presented a granular appearance or appeared clear. The cells were also distinguished from one another according as they possessed fine or coa.r.s.e granules. The granules are confined to the protoplasm of the cell, and it has been shown that they differ chemically, because their staining properties vary. Thus, some granules select an acid stain, and the cells containing them are then designated _acidophile_ or _eosinophile_;[1] other granules select a basic stain and are called _basophile_, while yet others prefer a neutral stain (_neutrophile_).

In human blood the following varieties of leucocytes may be distinguished:--

1. _The Polymorphonuclear Cell._--This possesses a nucleus of very complicated outline and a fair amount of protoplasm filled with numbers of fine granules which stain with eosin. They vary in size but are usually about 0.01 mm. in diameter. They are highly amoeboid and phagocytic, and form about 70% of the total number of leucocytes.

2. _The Coa.r.s.ely Granular Eosinophile Cell._--These large cells contain a number of well-defined granules which stain deeply with acid dyes. The nucleus is crescentic. The cells amount to about 2% of the total number of leucocytes, though the proportion varies considerably. They are actively amoeboid.

3. _The Lymphocyte._--This is the smallest leucocyte, being only about 0.0065 mm. in diameter. It has a large spherical nucleus with a small rim of clear protoplasm surrounding it. It forms from 15 to 40% of the number of leucocytes, and is less markedly amoeboid than the other varieties.

4. _The Hyaline_ (Gr. [Greek: hualinos], gla.s.sy, crystalline, [Greek: ualos], gla.s.s) _cell or macrocyte_ (Gr. [Greek: makros], long or large).--This is a cell similar to the last with a spherical, oval or indented nucleus, but it has much more protoplasm. It const.i.tutes about 4% of all the leucocytes and is highly amoeboid and phagocytic.

5. _The Basophile Cell_.--This possesses a spherical nucleus and the protoplasm contains a small number of granules staining deeply with basic dyes. It is rarely found in the blood of adults except in certain diseases.

_Functions._--These cells act as scavengers or as destroyers of living organisms that may have gained access to the tissue s.p.a.ces. They play an important part in the chemical processes underlying the phenomena of immunity, and some at least are of importance in starting the process of clotting.

They are constantly suffering destruction in the performance of their work. Many, too, are lost to the body by their pa.s.sage through the different mucous surfaces. Their origin is still obscure in many points.

The lymphocytes are derived from lymphoid tissue, wherever it exists in the different parts of the body. The polymorphonuclear and eosinophile cells are derived from the bone-marrow, each by division of specific mother cells located in that tissue. The macrocyte is believed by many to represent a further stage in the development of the lymphocyte. Their rate of formation may be influenced by a variety of conditions--for instance, they are found to vary in number according to the diet and also, to a considerable extent, in disease.

_Platelets._--The platelets or thrombocytes (Gr. [Greek: thrombos], clot) are the third cla.s.s of formed elements occurring in mammalian blood. There are still, however, many observers who consider that platelets are not present in the normal circulating blood, but only make their appearance after it has been shed or otherwise injured. They are minute lens-shaped structures, and may amount to as many as 800,000 per cubic mm. Under certain conditions, examination has shown that they are protoplasmic and amoeboid, and that each one contains a central body of different staining properties from the remainder of the structure. This has been regarded by some as a nucleus. On being brought into contact with a foreign surface they adhere to it firmly, very rapidly pa.s.sing through a number of phases resulting ultimately in the formation of granular debris. In shed blood they tend to collect into groups, and during clotting, fibrin filaments may be observed to shoot out from these clumps.

_Variations in the Blood of different Animals._--If we contrast the blood of different animals of the vertebrate cla.s.s we find striking differences both in microscopic appearances and in chemical properties.

In the first place, the corpuscles vary in amount and in kind. Thus, whilst in a mammal the corpuscles form 40 to 50% of the total volume of the blood, in the lower vertebrates the volume is much less, e.g. in frogs as low as 25% and in fishes even lower. The deficiency is chiefly in the red corpuscles, the ratio of white to red increasing as we examine the blood from animals lower in the scale. The corpuscles themselves are also found to vary, especially the red ones. In the mammal they are biconcave disks with bevelled edges, they do not contain a nucleus so that they are not cells. In the bird they are larger, ellipsoidal in shape and have a large nucleus in the centre of the cell.

In reptiles and amphibia the red corpuscles are also nucleated, but the _stroma_ portion containing the haemoglobin is arranged in a thickened annular part encircling the nucleus. When seen from the flat they are oval in section. In fishes the corpuscles show very much the same structure. A further very significant difference to be observed between the bloods of different vertebrates is in the amount of haemoglobin they contain; thus in the lower cla.s.ses, fishes and amphibia, not only is the number of red corpuscles small but the amount of haemoglobin each corpuscle contains is relatively low. The concentration of the haemoglobin in the corpuscles attains its maximum in the mammal and the bird. Since the haemoglobin is practically the same from whatever animal it is obtained and can only combine with the same amount of oxygen, the oxygen-capacity of the blood of any vertebrate is in direct proportion to the amount of haemoglobin it contains. Therefore we see that as we ascend the scale in the vertebrate series the oxygen-carrying capacity of the blood rises. This increase was a natural preliminary condition for the progress of evolution. In order that a more active animal might be developed the main essential was that the chemical processes of the cell should be carried out more rapidly, and as these processes are fundamentally oxidative, increased activity entails an increased rate of supply of oxygen. This latter has been brought about in the animal kingdom in two ways, first by an increase in the concentration of the haemoglobin of the blood effected by an increase both in the number of corpuscles and in the amount of haemoglobin contained in each, and secondly by an increase in the rate at which the blood has been made to pa.s.s through the tissues. In the lower vertebrates the blood pressure is low and the haemoglobin content of the blood is low, consequently both rate of blood-flow and oxygen-content are low. In contrast with this, in higher vertebrates the blood pressure is high and the haemoglobin content of the blood is high, consequently both rate of blood-flow and oxygen-content are high. We must a.s.sociate with this important step in evolution the means employed for the more rapid absorption of oxygen and for its increased rate of discharge to the tissues, the most important features of which are a diminution in the size of the corpuscle and the attainment of its peculiar shape, both resulting in the production of a relatively enormous corpuscular surface in a unit volume of blood.

Variations are also found in the white corpuscles as well as in the red, but these differences are not so striking and lie chiefly in unimportant details of structure of individual cells. Enormous variations are to be found in different species of mammals, but the cells generally conform to the types of secreting cells or phagocytes.

The platelets also differ in the different species. In the frog, for instance, many are spindle-shaped and contain a nucleus-like structure.

Birds' blood is stated to contain no platelets. The variations in number of these bodies have not been satisfactorily ascertained on account of the difficulties involved in any attempt to preserve them and to render them visible under the microscope.

Differences are also found in the chemical composition of the plasma.

The chief variation is in the amount of protein present, which attains its maximum concentration in birds and mammals, while in reptiles, amphibia and fishes it is much less. The bloods of the latter two cla.s.ses are much more watery than that of the mammal. Moreover, it has been proved that there are specific differences in the chemical nature of the various proteins present even between different varieties of mammals. Thus the ratio of the globulin fraction to the alb.u.min fraction may vary considerably, and again, one or other of the proteins may be quite specific for the animal from which it is derived.

_Clotting._--If a sample of blood be withdrawn from an animal, within a short time it undergoes a series of changes and becomes converted into a stiff jelly. It is said to _clot_. If the process is watched it is seen to start first from the surfaces where it is in contact with any foreign body; thence it extends through the blood until the whole ma.s.s sets solid. A short time elapses before this process commences--a time dependent upon two chief conditions, viz. the temperature at which the blood is kept and the extent of foreign surface with which it is brought into contact. Thus in a mammal the blood clots most quickly at a temperature a little above body temperature, while if the blood be cooled quickly the clotting is considerably delayed and in the case of some animals altogether prevented. For example, human blood kept at body temperature clots in three minutes, while if allowed to cool to room temperature the first sign of clotting may not make its appearance until eight minutes after its removal from the body. The process of clotting is also considerably accelerated by making the blood flow in a thin stream over a wide surface. The full completion of the process occupies some time if the blood be kept quiet, but ultimately the whole ma.s.s of the blood becomes converted into a solid. At this stage the containing vessel may be inverted without any drop of fluid escaping. A short time after this stage has been reached drops of a yellow fluid appear upon the surface and, increasing in size and number, run together to form a layer of fluid separated from the clot. This fluid is termed _serum_; its appearance is due to the contraction of the clot, which thus squeezes out the fluid from between its solid const.i.tuents. Contraction continues for about twenty-four hours, at the end of which time a large quant.i.ty (one-third or more of the total volume) of serum may have been separated. The clot contracts uniformly, thus preserving throughout the same general shape as that of the vessel in which the blood has been collected. Finally the clot swims freely in the serum which it has expressed.

The cause of the clot formation has been found to be the precipitation of a solid from the liquid plasma of the blood. This solid is in the form of very minute threads and hence is termed _fibrin_. The threads traverse the ma.s.s of blood in every possible direction, interlacing and thus confining in their meshes all the solid elements of the blood. Soon after their deposition they begin to contract, and as the meshwork they form is very minute they carry with them all the corpuscles of the blood. These with the fibrin form the shrunken clot.

If the rate at which blood clots be r.e.t.a.r.ded either by cooling or by some other process the corpuscles may have time to settle, partially or completely, in which case distinct layers may form. The lowermost of these contains chiefly the red corpuscles, the second layer may be grey owing to the high percentage of leucocytes present, while a third, marked by opalescence only, may be very rich in platelets. Above these a clear layer of fluid may be found. This is _plasma_. The formation of these layers depends solely upon the rate of sedimentation of these elements, the rate depending partly upon differences in specific gravity, and partly upon the tendency the corpuscles have to run into clumps. Horse's blood offers one of the best instances of the clumping of red corpuscles, and in this animal sedimentation of the red corpuscles is most rapid.

If now such a sedimented blood is allowed to clot the process is found to start in the middle two layers, i.e. in those containing the white corpuscles and platelets. From these layers it spreads through the rest of the liquid, being most r.e.t.a.r.ded, however, in the red corpuscle layer, and particularly so if the sedimentation has been very complete. Not only does the clotting process start from the layers containing the leucocytes and platelets, but in them it also proceeds more quickly.

These observations clearly indicate that the clotting process is initiated by some change starting from these elements.

The object of the clotting of the blood is quite clear. It is to prevent, as far as possible, any loss of blood when there is an injury to an animal's vessels. The shed blood becomes converted into a solid, and this, extending into the interior of the ruptured vessel, forms a plug and thus arrests the bleeding. It is found that clotting is especially accelerated whenever the blood touches a foreign tissue, for instance, the outer layers of a torn blood-vessel wall, muscle tissue, &c., i.e. in exactly those conditions in which rapid clotting becomes of the greatest importance. Yet another very pregnant fact in connexion with clotting is that if an animal be bled rapidly and the blood collected in successive samples it is found that those collected last clot most quickly. Hence the more excessive the haemorrhage in any case, the greater becomes the onset of the natural cure for the bleeding, viz.

clotting.

When we begin to inquire into the nature of clotting we have to determine in the first place whence the fibrin is derived. It has long been known that two chemical substances at least are requisite for its production. Thus certain fluids are known, e.g. some samples of hydrocele or pericardial fluid, which will not clot spontaneously, but will clot rapidly when a small quant.i.ty of serum or of an old blood-clot is added to it. The const.i.tuent substance which is present in the first-named fluids is known as fibrinogen, and that present in the serum or the clot is known as fibrin-ferment or _thrombin_.

Fibrinogen is present in living blood dissolved in the plasma; it is also present in such fluids as hydrocele or pericardial effusions, which, though capable of clotting, do not clot spontaneously. Thrombin, on the other hand, does not exist in living blood, but only makes its appearance there after blood is shed. It is not yet certain what is the nature of the final reaction between fibrinogen and thrombin. The possibilities are, that thrombin may act--(1) by acting upon fibrinogen, which it in some way converts into fibrin, (2) by uniting with fibrinogen to form fibrin, or (3) by yielding part of itself to the fibrinogen which thus becomes converted into fibrin. The experimental study of the rate of fibrin formation, when different strengths of thrombin solutions are allowed to act upon a fibrinogen solution, leads us to the probable conclusion that the first of these three possibilities is the correct one, and that thrombin therefore exerts a true ferment action upon fibrinogen. It is known that in the reaction, in addition to the formation of fibrin, yet another protein makes its appearance. This is known as fibrinoglobulin, and apparently it arises from the fibrinogen, so that the change would be one of cleavage into fibrin and fibrinoglobulin. It is very noteworthy that although the amount of fibrin formed during the clotting appears very bulky, yet the actual weight is extremely small, not more than 0.4 grms. from 100 cc.

of blood.