BLOOD AND OTHER STAINS.

BY

JAMES F. BABCOCK.

In many trials for homicide, especially in cases where the evidence is IN circumstantial, questions arise concerning spots or stains found upon clothing, weapons, furniture, carpets, walls, or other objects, and the scientific witness is expected to answer whether or not the stains are of blood or some other substance. If blood, are the stains old or comparatively recent? What was their origin? Are they human, or from some domestic animal? Are they stains of venous, arterial, or menstrual blood? Was the blood from a living or a dead body? Male or female? Adult or child?

To some of these questions answers may be given which are perfectly definite and reliable, but as to others it can only be replied that our present knowledge furnishes insufficient data for any certain conclusions.

In giving the results on these matters to which science leads us, we shall first briefly state the various physical and chemical properties of blood as it is found in man and animals, and then fully explain the application of these facts to the practical operations necessary for the solu tion, so far as possible, of the different questions we have stated.

CHEMICAL AND PHYSICAL PROPERTIES OF BLOOD.

Fresh blood is an opaque and somewhat viscous fluid slightly heavier than water. Its specific gravity, on the average, is in normal blood 1.055, but it is slightly less in women and children. In the higher animals the gravity is substantially the same as in man.

The color of blood varies from a bright scarlet to a deep purple, according as it flows from an artery or a vein. In very thin films as observed in the microscope, it is transparent and nearly colorless. The variations in color of venous and arterial blood are due to the degree of oxidation of the coloring matter, called hæmoglobin; hence, venous blood on exposure to the air becomes brighter, and arterial blood in certain diseases dependent upon a reduction in the supply of oxygen (asphyxia, etc.) is dark. On leaving the body the blood becomes gelatinous (coagulation), the change taking place in from three to fifteen minutes. Gradually, certain portions (coagulum) shrink in volume, and after a period of from twelve to forty hours there is a complete separation into thick red clots and a yellowish watery fluid (serum). The coagulation of the

blood may be hastened or retarded by a variety of circumstances. Moderate warmth accelerates, while cold retards coagulation. Access of air promotes coagulation, hence blood in thin layers thickens more rapidly than when it exposes a more limited surface. Coagulation takes place more readily when the blood flows upon rough surfaces; on cloth blood becomes clotted quicker than upon a smooth marble floor or polished furniture.

Chemical Composition of Blood.-1000 parts of blood contain, on the average, 795 parts of water and 205 parts of solids. The solids consist of albumen, fibrin, coloring matter containing iron, called hæmoglobin, cholesterin, and fatty bodies, various salts and extractive matters. The salts contain chlorin, sulphuric, phosphoric, lactic, oleic, stearic, uric, and hippuric acids, combined with potassium, sodium, calcium, and magnesium. The extractive matters contain small amounts of sugar, leucin, tyrosin, xanthin, creatin, and other substances. When examined by the microscope, blood is seen to consist of a colorless fluid (liquor sanguinis) in which are suspended large numbers of cell-like bodies called bloodcorpuscles.

One thousand parts of the liquor sanguinis and of the blood-corpuscles contain :

The solid constituents of each of these portions consist of:

The blood-corpuscles in their moist condition constitute about fifty percent. (47.2 to 54.2) of the total weight of the blood, and have a specific gravity of 1.088. The specific gravity of the liquor sanguinis is 1.028.

Of the various bodies entering into the composition of blood, there are only two which are of interest or importance to the medical jurist in his study of blood and blood-stains-hæmoglobin, which contains the coloring matter, and the blood-corpuscles. The chemical and spectroscopic phenomena produced by hæmoglobin afford positive evidence of the pres ence of blood, from whatever source it may have been derived; and the microscope, aided by the micrometric measurement of the diameters of the blood-corpuscles, gives all that may be determined concerning the origin of the blood, and whether it be human or otherwise.

Hæmoglobin.-The coloring matter of blood, now generally called hæmoglobin, was first described by Le Canu under the name of hæmatin. (Nouvelles Études Chimiques sur le Sang, Paris, 1852.) Stokes called it cruorin, and showed that it was capable of existing in two forms or states

of oxidation. Scarlet cruorin was the name given to the product found in arterial blood, and purple cruorin to that found in venous blood. (Stokes, Proc. Roy. Soc., 1864, p. 355.) Sorby described an intermediate body under the name of brown cruorin. (Sorby, Monthly Micros. Jour., London, vol. vi., p. 9.) Thudicum and Kingzett adopted the name hamato-crystallin. (Jour. Chem. Soc., London, September, 1876.)

When blood defibrinated by whipping is mixed with a 34-percent. solution of common salt, the corpuscles are gradually deposited, and the supernatant liquor may be decanted. On washing the deposit with a fresh portion of the salt solution the corpuscles are obtained free from serum. They consist of a stroma or colorless skeleton containing hæmoglobin, a little cholesterin, paraglobulin, fatty matters, and mineral salts. If the washed corpuscles are shaken with water and ether, the stroma, cholesterin, and fatty matters are taken up by the ether, while the coloring matter of the blood passes into solution in the water. On exposure to a low temperature a deposit of crystals is formed. They consist of hæmoglobin. One half its volume of alcohol may be added to the aqueous solution to promote crystallization. Hæmoglobin thus obtained consists of several proximate principles: an albuminous substance, which, when separated, is amorphous and colorless, and a crystalline body, called hamatin, having a formula C32H32FеN,Ó. (Kingzett.) Hæmoglobin is perfectly and freely soluble in water and dilute alcohol. By the action of acids or alkalies, or of any reagent capable of coagulating albumen, it is separated into hæmatin and the albuminous body above mentioned. The same change is produced by long exposure to the air, or by a shorter exposure to air containing considerable moisture or impurities such as are found in the atmosphere of cities. Exposure for a shorter period produces a brown substance intermediate between hæmoglobin and hæmatin, called met-hæmoglobin.

Hæmatin. Hæmatin is insoluble in water and ether, but is very slightly soluble in alcohol. It readily dissolves in ammonia water and in solutions of sodium and potassium hydrate. It is soluble in dilute acids, and especially in dilute citric acid. It is a very stable body, and when once formed may remain unchanged for years. If hæmatin or dried blood is heated with glacial acetic acid and a small amount of common salt, and the solution evaporated, a new combination is produced. It is generally considered to be hæmatin hydrochloride, but was named hamin by Teichmann, its discoverer, and it is by this name that it is generally called. It crystallizes readily from its solution in hot acetic acid. Hæmin crystals are insoluble in water, alcohol, ether, and dilute acids, but are sparingly soluble in ammonia water, and freely soluble in solution of sodium or potassium hydrate.

Recapitulation.-Fresh and unaltered blood yields crystals of hæmoOxidized blood or dried blood contains hæmatin.

globin.

Dried blood or blood treated with glacial acetic acid and salt yields hamin.

Fresh blood-stains are bright scarlet, and yield their coloring matter very readily to cold water. Hot water renders the stain more or less insoluble, on account of the coagulation of the albumen, while soap and water have a tendency to fix the color, from the conversion, in consequence of the presence of the alkali, of hæmoglobin into hæmatin.

Less recent stains are reddish brown or dark brown in color. They contain met-hæmoglobin. They yield but little of their coloring matter to water, while very old stains yield no coloring. Such stains are soluble in dilute citric acid, and give up their coloring to ammonia water.

Optical Properties of Blood-Coloring Matter.-When a solution of the coloring matter of blood is examined by the spectroscope, certain dark spaces called absorption-bands are observed. These bands in number and position vary according to the degree of oxidation of the bloodcoloring matter or the presence of reagents. The study of the absorptionbands under different conditions has led to the discovery of a method (spectrum analysis) which may be relied upon with absolute certainty for the identification of blood and for distinguishing it from all other substances. A brief description of the principles involved in this method and the apparatus employed for the purpose is here given.

Spectrum Analysis and the Spectroscope.-When a beam of light is passed through a narrow slit of 1-100 to 1-1000 of an inch in width, and then through a prism and allowed to fall upon a white screen, an elongated colored image is produced containing all the brilliant hues of the rainbow.

If the light be passed through several prisms no additional colors are produced, but the image is lengthened and the colors more widely separated. This image is called the spectrum. If it be observed through a magnifying-lens or a small telescope, it is found to be crossed at right angles by numerous dark lines. These lines were first carefully observed by Fraunhofer in 1815, and have since been called Fraunhofer's lines. This observer found that the lines always kept their position, provided the same prism and lenses were employed, and he made a map or chart of them. He selected eight of those which appeared wider than the others, and named them by the letters of the alphabet from A to H. These lines have been adopted as standards of comparison for denoting the position of any set of colored rays which may be submitted to examination. Lines A and B are in the red portion of the spectrum; C, in the red, near the orange; D, between the yellow and the orange; E, in the green; F, on the borders of the green and blue; G, in the dark blue; and H, at the extreme end of the violet. Kirchoff in 1859 proved that Fraunhofer's lines were due to the presence of certain gases in the solar atmosphere which have the power of absorbing the same rays of light as those emitted by the heated body producing them. Later it was found that various colored solutions had a similar property, so that light passed through them produces a spectrum crossed by dark bands (absorptionbands), which vary in position and intensity according to the nature of the substance or its strength of solution. An instrument adapted to the examination and study of the spectrum or its absorption-bands is called a spectroscope. The ordinary form of instrument such as is used in laboratories for the analysis of colored flames is not adapted to the examination of the absorption-bands of blood, although it may be used in the absence of one specially designed for this purpose, called the microspectroscope, and sometimes described as the spectrum-microscope. It consists of a series of prisms so arranged that they may be attached to the microscope either above the eye-piece or in the ordinary position of the objectglass. The best forms are provided with a scale, which enables the exact position of the bands to be determined, and a supplementary stage, by which the spectrum of one body may be compared with another, the two

spectra being visible side by side at the same time. Sorby in 1866 suggested the form of instrument generally used. Its essential features are shown in the accompanying outline.

Two rectangular prisms of flint-glass are separated from each other by a prism of crown-glass, and two other similar prisms are attached one on each end of the combination. These are cemented with Canada balsam. This compound prism is mounted in a tube, F, having a cap with an elongated opening at A and a circular stop at B. The tube is constructed so that it may be slipped over the upper lens of the eye-piece. The upper lens, G, is compound and achromatic, and is mounted so that the focus may be adjusted by suitable rackwork or by turning the milled head, H. At I is a slit capable of being adjusted to a wider or narrower

opening, and a right-angled prism, C, is fixed half-way over it. By this means light passing through an opening at E is reflected through half of the slit, while light coming through the field-glass from the object passes through the other half. In this way may be seen side by side the spectrum of the light passing from the object under examination and that produced by the light coming from the stage E, which holds the standard for comparison. The supplementary stage has an adjustable slit by which the two spectra may be made to appear of equal brilliancy. The solution or object to be examined is placed upon the stage of the miscroscope and strongly illuminated by the mirror; the standard for comparison is contained in a sealed tube held by springs on the stage E.

Fig. 10 represents the spectroscopic eye-piece made by Zeiss. In this