Monthly microscopical journal: transactions of the Royal Microscopical Society, and record of histological research at home and abroad
Royal Microscopical Society (Great Britain)
= s ‘a > s Ss : s s « : s < s s A. s s < s « < 4 Fao Toray Oe EW YORK BOTANICAL he My fa Besa " a 5 ==) - ES a IE 7: ZAcetiece)) Gartner D*. ooo ee eee Yee —=—>>>= ry = a aaa. 4 THE MONTHLY -MICROSCOPICAL JOURNAL: TRANSACTIONS ROYAL MICROSCOPICAL SOCIETY, RECORD OF HISTOLOGICAL RESEARCH AT HOME AND ABROAD. EDITED BY HENRY LAWSON, M.D., F.R.MS.. Assistant Physician to, and Lecturer on Physiology in, St. Mary’s Hospital. VOLUME VI. soos 1BRAR ae \ 2 BREW YORK BOTANICAL Carpe LONDON: ROBERT HARDWICKE, 192, PICCADILLY, W. MDCCCLXXI, me LONDON: PRINTED BY WILLIAM CLOWES AND SONS, STAMFORD STREET AND CHARING CROSS. “AMI Baas BS ae ie - WiWest &Co.se. DEG Tt I9Ul g m3GP r BY YOO K z rah: Seal THE MONTHLY MICROSCOPICAL JOURNAL. JOLY 1,. 1871, I.—On Bog Mosses. By R. Brarrnwarre, M.D., F.LS. (Read before the Royau Microscopicau Society, June 7, 1871.) Part I. For a considerable time the plants known as bog mosses have attracted attention, not only by the masses in which they are found growing, but by their peculiarity of structure, and the difficulty of finding characters by which to establish the species, for the varieties are endless, and such a common facies is impressed on the whole group that Linneus regarded them all as one species, which he named Sphagnum palustre. In our own time, however, the careful use of the microscope has revealed to us their wonderful organiza- tion, and enabled us to establish many species. Up to the publication of Professor Schimper’s magnificent trea- tise,* the bog mosses had been universally associated with the other mosses in one class, but in that work they are placed apart, and EXPLANATION OF PLATE XC. Fic, 1.—Prothallium with young plant. 2.—Sphagnum cymbifolium. Vertical section of stem passing also through two leaves, and the base of a branch fascicle. » 3.—Ditto. Transverse section. These show the pith, the woody cylinder, and the four layers of bark cells. » 4—Cells of pith. 5, 0—Ditto of wood. », 6.—Ditto of bark. » 7—Cells of a branch leaf of S. cymbifolium. », 72.—Transverse section of a leaf of S. squarrosum. » 8—Cells of a branch leaf of S. acutifolium, seen from the back. », 8x.—Transverse section of same. », 9.—Male flower catkin of S. cymbifolium. », 10.—Ripe antheridium with paraphyses. ,, 11.—Vertical section of a capsule still enclosed in the calyptra. c, calyptra ; s, cavity of sporangium; p, pedicel of capsule enveloped by the vaginula, (Fig. 1 from a specimen lent by Mr. Howse, the rest from Schimper’s work.) * ¢Rntwickelungs-geschichte der Torfmoose,’ 1858. VOL. VI. B 2 _ Transactions of the hold equal rank with Mosses and Hepatic; the three classes, Bryinz, or frondose mosses, Sphagnine, or bog mosses, and Hepa- ticine, or liverworts (the Laubmoose, Torfmoose, and Lebermoose of the Germans), thus forming one great muscal alliance. As it may be of interest to know something of the histology of the Sphagnums, and except the late Mr. Wilson’s admirable sketch of the family in ‘ Bryologia Brit.’ we have little in our own language bearing on the subject, I purpose on this occasion to point out the chief characters which distinguish them from mosses, and at a future time, with your permission, give an account of all the British species, with illustrative figures of their structure. Commencing with the spore, we find that on germination it does not produce the much-branched confervoid prothallium of mosses, but if growing on the wet peat, a lobed foliaceous production results, exactly like one of the frondose Hepatice, as Anthoceros, and from this the young plants grow. This was first observed by Hofmeister in 1854. If germination occur in water, the prothallium is a fine filament, the lower end of which forms roots and the upper enlarges into a nodule from which is developed the young plant. As-soon as branches form on the plants, in both cases the prothallium and roots wither away. Next, the stem, which in mosses consists of uni- form texture throughout, is in Sphagnums much more differentiated, for we observe—Ist, a medulla, or central pith, of long cylindric cells by which a current of sap is constantly passing to the growing apex ; 2nd, a woody cylinder of firmer prosenchymatous cells; and 3rd, an outer, or bark-layer of one to four strata of thin walled cells, always larger than the rest, and which in S. eymbifoliwm alone contain spiral threads and foramina. The branches also are quite pee and differ altogether in arrangement from those of mosses. n the young state they are crowded into a capitulum at the top of the stem, and as the internodes elongate they become separated, and are then seen to be in lateral fascicles of three to ten, the bundle originating at one side of a leaf base, every fourth leaf in the spiral giving rise to a branch fascicle. Of these branches, one part spread out horizontally and then arch down, the rest being attenuated, pendulous, and pressed back to the stem, and by these water is conducted to the very apex; in- deed, Professor Schimper compares their action to a hydraulic pump, fora tuft of plants placed dry in a flask of water immediately carry on the fluid by the lower bark cells, and empty the flask by a discharge of drops from the down-bent capitulum, but if some of the branches and bark be removed, no passage of fluid takes place beyond the injured part. Of the sponge-like retention of moisture by Sphagnums, we have often unpleasant experience when stepping on the green turf-like surface of its beds, only to plunge deep into their interior, and Royal Microscopical Society, 3 emerge soaked with the water concealed therein. They can also absorb atmospheric moisture and transmit it downward, thus taking up again at night what they had lost during the day ; and by this perpetual interchange the stagnant pools in which they flourish never become putrid. ‘The bark of the branches consists only of a single layer of cells in communication with the innermost layer of the stem bark ; but these are of two forms, for besides the ordinary kind, there are others of a flask shape, larger, and with a circular orifice, one of which always falls at a leaf-insertion. The leaves of bog mosses are very peculiar, and are well known as an elegant microscopic object ; those of the stem are more remote from each other than in mosses, and in all the species two complete spirals contain five leaves (phyllotaxy 2); their form is ovate or tongue-shaped, with the base frequently more or less auricled. Those of the branches are much narrower, and not only vary on the two forms of branch, but on different parts of each. If we look at a Sphagnum leaf in fluid with a sufficient power, the first thing that strikes us is the beautiful sigmoid form of the areolation, or cellular network; and next, the presence of a delicate fibril forming spirals or rings on the inner wall of each cell, and by which the thin membrane is kept expanded, while perforating the mem- brane are distinct apertures or pores through which it is common enough to find infusoria have passed, which may be seen sporting about in the cell cavity ; and thirdly, that these large prosenchyma- tous cells are always void of chlorophyl, and hence want the lively green colour so noticeable in true mosses. _ This, however, is not all. A more careful examination will show that between the walls of these hyaline cells are placed extremely narrow parenchymatous cells, which do contain chloro- phyl. Moldenhawer first detected these in 1812; yet Carl Miiller, in his ‘Synopsis Muscorum, terms them intercellular ducts; in no case is there any approach to the formation of a nerve or midrib. According to the colour of the chlorophyl in these parenchym cells, is the colour of the Sphagnum tuft, only seen, indeed, in the living or moist state, but presenting endless shades of rosy red, purple, vinous red, bluish green, olivaceous, apple-green, and straw colour. When dry or dead, the hyaline cells lose their transparency, and all the species become more or less a dirty white. The spiral threads are nothing but thickenings on the inner wall of the cell membrane, such as occur frequently in the tissues of phzenogamous plants; in one species only are they altogether absent, viz. the American S. macrophyllum, which Professor Lindberg separates as the genus Tsocladus. The male flowers of Sphagnums differ also from those of mosses, and in their arrangement, and the form of their antheridia, resemble those of Hepatice; they are grouped in catkins at the tips ¢ lateral B + Transactions of the branches each of the imbricated perigonial leaves enclosing a single slobose antheridium on a slender pedicel, which is inserted at the side of the leaf base. Paraphyses surround them, but instead of being simple, as in mosses, they are very long, much branched, and of cobweb-like tenuity. The perigynium of the female flower-sheath terminates one of the short lateral branchlets at the side of the capitulum, and is of a deep-green colour, with large sheathing leaves; the archegonia do not differ from those of mosses. After impregnation the fruit receptacle enlarges and extends itself into a pseudopodium, on which the capsule is sessile; the vaginula was thought to be absent from these plants, and hence Bridel formed for them a section “ Musci evaginulati.” It is, how- ever, very distinct, being the disk-shaped enlargement at the apex of the receptacle in which the expanded bulbous rudiment of the pedicel of the capsule is completely buried. The calyptra, or outer cell-layer of the archegonium, has not the definite form common to mosses, but is a very thin colourless membrane closely investing the capsule, by the enlargement of which it is torn irregularly into shreds, the lower portion being left attached to the vaginula. The capsule up to maturity remains in the perichetium, but after that the receptacle elongates and carries it upward on this pseudopodium, which does not correspond to the pedicel of a moss, that being always found above the vaginula; still, a pedicel does exist in Sphagnum, but it is bulb-shaped, and enveloped by the vaginula. ‘The capsule itself is very uniform in all the species, being almost spherical, the lid only slightly convex, without any beak or point, and we never find any trace of a peristome. The sporangium, or spore sac, does not attain the development found in mosses, for it appears like a hollow hemisphere in the interior of the fruit, its outer wall bemg coherent with the inner cell-layer of the capsule, its inner wall with the columella lying beneath it. The spores somewhat resemble those of Lycopods in being of two kinds—macrospores produced by fours in a mother cell and tetrahedral in form, and microspores which are more spherical, and not half the size.. With respect to the function of these plants in the formation of peat, I cannot do better than quote Professor Schimper’s words. He says :—“ Unless there were bog mosses, many a bare mountain ridge, many a high valley of the temperate zone, and large tracts of the northern plains, would present an uniform watery flat, instead of a covering of flowering plants or shady woods. For just as the Sphagna suck up the atmospheric moisture, and convey it to the earth, do they also contribute to it by pumping up to the surface of the tufts formed by them the standing water which was their cradle, Royal Microscopical Society. 5 diminish it by promoting evaporation, and finally, also by their own detritus, and by that of the numerous other bog plants to which they serve as a support, remove it entirely, and thus bring about their own destruction. Then, as soon as the plant detritus formed in this manner has elevated itself above the surface water, it is fami- har to us by the name of turf, becomes material for fuel, and all Sphagnum vegetation ceases.” Little noticed, then, as these plants may be, they perform func- tions in the economy of nature which cannot be overlooked ; and it is for the purpose of inviting more general observation of their structure that I bring forward these remarks. Some seventeen British species are known, all of which I hope to illustrate; and in carrying out this object, I shall feel thankful for good specimens of any of our species, but especially those occurring in the north of Scotland, for the wild moorlands of Northern Europe have so far proved to be the most prolific in new species. ey Il.—Structure of Podura Scales. By F. H. Wenuaw, Vice-President R.M.S. Mr. Molnrire has kindly presented me with a number of slides ; two of them are superb specimens of the test Podura, L. ewrvicollis, with markings remarkably dark and distinct. In one of the best scales there is a singular case of fracture, evidently caused by the slipping of the cover, which has dragged the specimen asunder near one-third from the upper end, which is separated to a short dis- tance. Projecting from the largest portion is a fragment, partly torn away, as shown by the accompanying cut, \ \ which i ae outline ; NSS sketch taken by the ( SS camera lucida, 4000 N — diameters. On the SSS right-hand extremity the longitudinal tear has taken place close to a rib, or marking, which is nearly iso- 4,090 lated. The transverse tear at the bottom leaves most of the ends of the ribs exposed and projecting, as the membrane has torn away from behind them. The extremities of ribs 4 and 5 (counted from the right) are particularly plain and prominent. I am willing to submit this specimen to the inspection of anyone doubting the fidelity of the tracing. How it can be maintained or admitted that these waved or con- stricted ribs which give rise to the “ note'of exclamation” markings are “illusory,” I am at a loss to imagine.’ The question can only be determined by fragmentary pieces; but in the Podwre, from the toughness and absence of brittleness in the scales, these are very difficult to obtain, and it is seldom that a fortunate accident occurs in the way shown. It was by fragments that the structure of the diatoms became known, wherein the silicious nodules could be traced down to a single atom, and their form is not now a question of dispute. Mr. Hennah, and others, have demonstrated how a number of different patterns, not the least indicative of real structure, may be produced by transparent bodies of regular form, such as glass rods and bosses, &c. Much more incomprehensible would these appear- ances be if the transparent ribbings were twisted, lobated, or in the peculiar form of a Podura marking. The patterns would be innu- merable with oblique light, and the whole area might be made to Structure of Podura Scales. t consist apparently of ‘‘ illusory beads,” and the belief now appears to be, with the most unbiassed observers, that they are such. The Degeeria domestica, or speckled Podura, when shown opaquely * under a 7'sth or upwards, is a specially beautiful object. The scales are apparently much thicker than in other species, and the ribbings or !!! markings are of a reddish-brown colour—not beaded, but slightly constricted at regular intervals, like the short antennee of some insects, and in the deep intercostal spaces there are numerous thin septe, or transverse bars, very fine and distinct, of a greyish tint. Both these and the slightly “ varicose” spaces on the ribs may be displayed in the form of beads, by dodging the illumination. Where practicable, some form of opaque illumination should always be employed for verifymg the structure of these objects, for we are im this case quite free from the errors of diffraction, which more or less accompany objects seen by trans- mitted light, and cause an indistinctness of outline. * The only plan at present known to me of illuminating these and other dry test-objects is the one that I recently described, and consists in mounting them, not on the cover, but on the slide itself. When the light is totally reflected from - the upper surface of this at the part where the object is in contact, total re- flexion will not occur, but the light will pass into the object, and in many cases— as with some diatoms and the Podura—illuminate it with sufficient brilliancy on a black field to be plainly seen with the highest powers and eye-pieces. There are several methods of securing this total reflexion, some of which are described in my paper “On a Method of Dluminating Opaque Objects under the Highest Powers of the Microscope,” read before the Microscopical Society on March 26th, 1856, and published in the ‘ Transactions.’ One of them consists of a solid glass parabola with a flat top, upon which the slides were placed, with an intervening film of highly refractive medium ; this was made at the time, and ever since has been in use, mostly for viewing aquatic animalcules and living diatoms, &e., for which it is especially convenient. The parabola is first lowered so as to bring its top below the level of the brass table, and the object in a drop of water is covered with thin glass. The parabola is then screwed up till the object is secured. The rotifers are beautifully shown this way; but I have found that for objects on slides its use is not so convenient as the small truncated or nearly hemispherical lens and the ordinary parabola, as this arrangement affords greater facility for traversing the object, as there is sufficient play for the lens for this purpose in the hollow top. As some have complained of a difficulty in using this, I repeat the directions. The little lens is first patched on under the slide with a minute drop of oil of cloves or turpentine where the Podura scales or other objects lay thickest. The slide is then put on the stage with the lens in the top of the parabola. The next thing is particularly to obtain parallel light. A large bull’s-eye condenser should be employed with its flat side towards the lamp: hold a sheet of paper over the plane mirror, and see that the light falls in the centre, and is of the same area as the bull’s-eye or about fills the mirror; then go through the ordinary adjustments, and if the field is not quite black raise the stop of the parabola: look at the objects with a low power, those only that are brilliantly luminous are on the slide. Select a proper one, and if it does not appear nearly in the centre of under lens, shift this beneath it. One cause of failure may be the absence of objects detached from the cover on the slide; but in English mounted objects, either of diatoms or scales, I have in most cases found a few straggling specimens that have left the cover. Possibly the figure of some of the paraboloids may have degenerated since I sup- plied the original steel template near twenty years ago, which must have been worn out before now; but this I will see to. ( 8 ) III.—On some New Parasites. By T. Granam Ponton, F.ZS., Honorary Secretary Bristol Microscopical Society. PuatE XCI. Some time ago I described a new parasite from the tiger, in the pages of this Journal. I now propose to present to its readers four parasites, which I believe to be hitherto undescribed, or new. Menopon ptilorhynchi (mihi) (Plate XCL, Fig. 1).—Colour bright fulvous. Head obtusely subtriangular; clypeus rotundate, vertex rounded, base concave. Two broad irregular chestnut mark- ings extend from the insertion of the antennz to the eyes, which are connected at that point by a semilunar chestnut line, a chestnut spot in the centre of the clypeus; prothorax elliptical ; metathorax transverse ; abdomen ovate, hairy ; all the segments, except the last, have a chestnut spot; legs long, tarsi clavate. Length 2°115 mil. Habitat, Pélorhynchus holosericeus. Nirmus Nitzschit (mihi) (Plate XCI., Fig. 2).—This species is probably the same as that mentioned in Giebel’s list of the Halle Collection, without either name or description. Supposing this to be so, and that it is undescribed and unnamed, I have ventured to call it Nitzschit, after the indefatigable student of these peculiar insects, C. L. Nitzsch, and beg to give the following description of it :—Colour pale yellowish-white. Head panduriform, clypeus rounded, antennz rather long, second joint longest. Prothorax not so wide as the head ; metathorax oblong, trapeziform. Abdomen lanceolate, a long fascicule of hair between each of the four last segments. Legs somewhat clavate. Length 2'538 mil. Habitat, Péclorhynchus holosericeus. Docophorus Dennyti (mih) (Plate XCI., Fig. 3).—This species I have dedicated to the late Mr. Denny, of Leeds, to whom I am indebted for much kind advice and assistance in the study of the Anopleura. Colour tawny. Head triangular, clypeus produced entire; trabecule large, broadly truncated; antennz rather long. Clypeus bordered by a chestnut line, with a transverse semilunar marking of the same colour, a similar one on the occiput; a broad irregular chestnut mark extends from the eyes to the prothorax. Prothorax transverse, angles rounded, metathorax transverse. Abdo- men ovate, hairy; pale fulvous, with a chestnut border. Length, 3:173 mil. Habitat, Prismites Mexicanus. Trichodectes leporis (mihi) (Plate XCI., Fig. 4).—Colour bright fulyous yellow, a dark chestnut spot at the eyes connected by a diagonal line with a line of the same colour on the occiput. Head suborbicular ; clypeus rounded, vertex convex, lateral margin deeply sinuated ; eyes prominent ; antenn small, last joint broadly clavate ; prothorax transverse ; metathorax not so wide as the head. Abdo- men ovate, fulvous, hairy. Tibize clavate. Length, 2-538 mil. Habitat, Lepus cannabinus, Curton, May, 1871. Sy Wi) J ‘ e ade") IV.—On some Improvements in the Spectrum Method of Detecting Blood. By H.C. Sorsy, F.R.S., &e. In the following paper I shall give a condensed account of what I have been able to learn in connection with this subject, and omit everything that does not bear directly on determining whether any stain is, or is not, due to blood. There does not appear to be any probability of our being able to decide by this means whether it is, or is not, human. The spectrum-microscope used in these inquiries should have a compound prism, with enough, but not too great, dispersive power, or else the bands would be as it were diluted, and made less dis- tinct. A combination of two rectangular prisms of crown glass, with a rectangular of very dense flint, and another of less dense, of such an angle as to give direct vision, turned towards the slit, as lately made for me by Mr. Browning, appears to be the proper medium, and has other important advantages. The cells used for the experiments should be made from barometer tubing, and be about one-eighth of an inch in internal diameter, and half an inch long, one end being fastened to a piece of plate glass with purified gutta-percha, like an ordinary cell for mounting objects in liquids. It is, however, a very great advantage to insert between the plate and the cell a diaphragm of platinum foil, having a circular hole about two-thirds of the internal diameter of the tube, fixed so that its centre corresponds with that of the cell. This prevents any light passing upwards that has not penetrated through the whole length of the solution, which is very important when using direct concentrated sunlight to penetrate through turbid or very opaque liquids. A small spatula made of stout platinum wire, flattened at the end, is very convenient for adding small quantities of the re- agents ; and they should be stirred up in the cells with a platinum wire, flattened and turned up square at the end, like a small hoe. The reagents commonly employed are a somewhat diluted solution of ammonia, citric acid, the double tartrate of potash and soda, used to prevent the precipitation of oxide of iron, and the double sul- phate of the protoxide of iron and ammonia, employed to deoxidize ; but in some special cases diluted hydrochloric acid, carefully-purified boric acid, and sulphite of soda are required. The character of a stain varies much with its age, and with the nature of the substance on which it occurs. If quite recent, and if the substance has no immediate influence on blood, the stain would contain little or no colouring matter but hemoglobin. This is easily dissolved by water, and when properly diluted—neither too strong, nor too weak—it gives the well-known spectrum, with two dark absorption-bands in the green. The addition of a very little 10 On some Improvements in the ammonia and a small quantity of the double tartrate produces no change, but on adding a small piece of the ferrous salt, about oth of an inch in diameter, and carefully stirring, so as to mix without much exposure to the air, these bands gradually fade, and are replaced by the single broad and fainter band of deoxidized haemoglobin. When stirred up so as to expose well to the air, the two original bands of oxidized hemoglobin can be seen again. On gradually adding a little citric acid, until the colour begins to change, these bands slowly fade away; and, if the amount of blood was consi- derable, a faint band would make its appearance in the red. When previously deoxidized, this solution may be turbid, but not so as to interfere with the result. The addition of excess of ammonia makes all clear again, but does not restore the original bands, or only to a slight degree, thus showing that a permanent change is produced by citric acid—the hemoglobin is changed into hematin. This alone serves to distinguish blood from by far the greater number of coloured substances, which after being changed by acid, are restored by alkalis to the original state. On deoxidizing with the ferrous salt, we obtain the well-marked spectrum of deoxidized hematin, with one very dark and another much fainter band in the green, almost or quite invisible when the quantity is small. If too much citric acid or double tartrate had been added, this solution might be turbid; but, if all had been properly managed, it would be quite clear. Since the deoxidization takes place rather slowly, especially in cold weather, it is well to slightly stir up the ferrous salt at the bottom, completely fill up the cell, cover it with a piece of thin glass, remove the excess of liquid with blotting paper, and mix the solution by turning the tube upside down, over and over again. On reoxidizing the solution by stirring, the bands of deoxidized heematin disappear, and the two bands of hemoglobin will probably be recog- nized, owing to citric acid not changing the original merely into hematin, but also giving rise to some methemoglobin. The whole of these facts may be seen with a single cell, containing about 7)oth of a grain of blood, and any experimenter should become quite familiar with them before applying this method to suspected stains in cases of importance. Very faint bands are best seen by lamplight. On exposure to the air in a damp place, a blood-stain may be completely decomposed by the growth of mould, but when not thus destroyed it is partly altered into hematin. If, however, kept dry, the hemoglobin gradually changes into a variable mixture of methe- moglobin, hematin, and a brown substance not yet much studied. This change takes place far more rapidly in the acid atmosphere of towns and houses, especially when gas 1s burned, than in the open country ; but it does occur even in the purest air, and in glass tubes hermetically sealed. The presence of a weak acid in perspiration may also cause a stain on a worn garment to be completely changed Spectrum Method of Detecting Blood. 11 in a very short time, and the presence of a stronger acid on dirty clothes may at once alter the hemoglobin into hematin. On digesting in water a stain that has been kept until all the hemoglobin has disappeared, the methemoglobin dissolves. When the solution is sufficiently strong, this shows a band in the red, and two fainter in the green. The addition of ammonia removes that in the red, makes those in the green much darker, and develops a spe- cial very narrow band in the orange. When deoxidized this solution gives deoxidized hemoglobin. Since methemoglobin is formed at once from hemoglobin by the action of a great number of different oxidizing reagents, and since it can be reconverted into oxidized hemoglobin by slight deoxidization, I am inclined to look upon it as a peculiar oxidized modification. On adding a little of the double tartrate and of the ferrous salt to even a dilute solution from an old stain, the methemoglobin is deoxidized, and the well-marked spec- trum of fresh blood can be seen. If left too long, the spectrum of deoxidized hemoglobin is developed, but on well stirring, that of the oxidized reappears, and the various other spectra may afterwards be obtained, as described above. That part of the stain, insoluble in water, which is chiefly hematin, may be dissolved in dilute citric acid or ammonia, and when deoxidized the spectrum seen to even greater advantage than when fresh blood is employed, because there is no general shading in the green, due to there having been methemoglobin mixed with the hematin. We may thus obtain an excellent spectrum from a blood-stain nearly fifty years old. In very old stains all the methemoglobin has disappeared, and some- times even a considerable part of the hematin has been altered into another brown colourmg matter, which does not give any well- marked spectrum. When a blood-stain has been made sufficiently hot to coagulate the albumen, neither water, citric acid, nor cold ammonia will dis- solve it, but by heating in dilute ammonia the hematin is easily dissolved, and may be detected either before or after concentrating the solution by evaporation. I may here say that the spectrum of deoxidized hematin can in no way be better seen than by deoxidizing a solution of fresh blood that has been boiled with dilute ammonia, which gives rise to a very pure hematin. In applying these principles to the detection of suspected stains, it is desirable in the first place to examine a portion of the unstained fabric, to ascertain whether any colour is dissolved from it by water, and whether the solution has an acid, or alkaline, reaction. It-is also important to ascertain whether colour is dissolved from the fabric by dilute citric acid or dilute ammonia, and if so, to deter- mine whether this would in any way interfere with the recognition of blood by the processes described above. In the case of scarlet cloth and of some other red fabrics, much colour is dissolved out by 12 On some Improvements in the ammonia, but not by citric acid, which ought therefore to be used, whereas in other cases ammonia is the best solvent. Unless the stain is faint, a portion should be soaked in a few drops of water in a watch-glass, the liquid squeezed out, allowed to stand a short time in the glass, so as to deposit any small portions of the fabric, and poured into one of the experiment cells. If the stain had been recently made, and had not been changed by any special action, a solution of hemoglobin would be obtained, and the various spectra could be seen one after the other, as already described. If, however, the stain were a few days or a few weeks old, we should obtain a mixture of hemoglobin and methemoglobin, or the latter alone. The various spectra could then be developed, and compared side by side with those from fresh blood, to be sure that there is complete correspondence in the position and relative intensity of the bands. The residue insoluble in water should then be dissolved in dilute citric acid or ammonia, according to the nature of the fabric, and the spectrum of deoxidized hematin developed. If insoluble in cold citric acid or ammonia, hot ammonia should be tried, since the stain might have been so heated as to coagulate the albumen. If it be desirable to keep the specimen of deoxidized hzematin for subsequent reference, the cell may be covered with a piece of thin glass, and, after removing the excess of liquid, the edge of the cover painted round with gold-size. When properly managed, such an object will show a perfectly good spectrum, even after many weeks. If therefore we have a sufficient amount of a moderately old stain, we may easily see in succession the seven very different spectra of the following solutions:—1. Neutral methemoglobin. 2. Alkaline methemoglobin. 3. Deoxidized hemoglobin. 4. Oxi- - dized hemoglobin. 5. Acid hematin. 6. Alkaline hematin. 7. Deoxidized hematin. If the amount was very small, only Nos. 4 and 7 would show distinct bands, and the rest would be characterized rather by their comparative absence; and it must always be borne in mind that Nos. 1 and 2 may be modified by the presence of unaltered hemoglobin, No. 3 by that of dissolved hematin, and Nos. 5, 6, and 7 by that of undecomposed heemo- globin or methemoglobin. It would be easy to obtain other preparations, and to see several other spectra derived from blood, but it appears to me unneces- sary, since the above are so remarkable and unique in the manner in which they are produced, one after the other, especially by deoxidization and reoxidization on stirring, which seldom occurs in other colouring matters, that they afford as satisfactory a test for blood as could be desired, and still more so when we consider not only the general character of the spectra, but also the exact position of the absorption-bands, and that some are so most unusually dis- tinct. Spectrum Method of Detecting Blood. 13 The above directions apply to simple cases, where the amount of material at command is amply sufficient, and the fabric on which the stain is found does not contain anything that makes the blood insoluble, or interferes with the various tests. I shall, however, now describe what should be done in cases which are made specially difficult by various causes. If the stain were very faint, from the presence of very little blood, or if the greater part had been re- moved by washing with water, it might be desirable not to divide the material, but to examine the whole at once. The stained por- tion should therefore be digested in a few drops of dilute citric acid or ammonia, and the presence of hematin determined, as already described. If faint and spread over a considerable surface, it might be well to digest in citric acid or ammonia diluted with much more water than would fill the experiment cell, and afterwards concen- trate the solution by gentle evaporation. By this means blood could be detected, even when considerable effort had been made to remove it, and only a faint brown tinge left, just visible on white linen. ‘There would generally be no difficulty in the case of a stain on cloth which had been sponged, for enough blood solution would be left in the fabric. The presence of mordants in cloth or prints may require us to somewhat modify our proceedings, especially if the stain had been made wet, and to a great extent removed, so that we have only the dried-up solution of blood, thoroughly incorporated with the mor- dant. Certain kinds of brown cloth are of such a character, and about seven years ago portions of a wetted stain were sent by me to a number of the highest authorities in the detection of blood, and they said that neither they nor anyone else could recognize it. However, by proper care, I found that after a lapse of six years it could be detected by the spectrum method. The best plan was to digest a portion of the cloth in dilute ammonia, and to squeeze it well over and over again, with a pair of forceps, and finally with the finger and thumb, so as to obtain as much of the solution as possible. This was very turbid, but when deoxidized in the usual manner, and illuminated by concentrated light direct from the sun itself, the band of deoxidized hematin was quite distinct. When the cell was kept for a while, so that the insoluble part settled to the side, no band was visible, and therefore the hematin was eyi- dently combined with the mordant. It will thus be seen that it may be most important not to filter or allow the insoluble matter to subside, but to overcome the opacity by means of a sufliciently intense light. If the sun could not be made use of, the lime or electric light would no doubt be the best substitute. When fresh blood solution is agitated in a test tube with vege- table soil, and left until quite clear, the colouring matter is com- pletely carried down with the earth. Dilute ammonia, however, 14 On some Improvements in the dissolves out hematin, and therefore, in testing portions of soil, they should be digested in considerably more of that solvent than will fill an experiment cell, and after the solution has become quite clear it should be concentrated by evaporation. The spectrum of deoxidized hematin may then be seen by following the ordinary method. The same process should be adopted in examining stains on clothes impregnated with earth or earthy dust, and marks on iron contaminated with much rust, if water will not dissolve out unaltered blood or methzemoglobin. The importance of being able to detect blood-stains on leather was prominently brought before me by a case in which the trial of a suspected person depended on the nature of certain dark marks on his gaiters. The presence of tannic acid so completely mordants the blood, that neither water nor citric acid will dissolve it, and am- monia gives rise to a most inconveniently dark solution. If the stain is on the surface, and has never been wetted, a thin shaving should be cut off, so as to have as much blood and as little leather as possible, and the blood should be dissolved off without exposing the solution to the action of the leather itself. This may be accom- plished by taking one of the experiment cells, nearly filled with water, bending the shaving, and inserting it into the upper part of the tube, so as to touch the water, being careful to arrange it so that the stain may be on the convex side of the leather, and in contact with the water. When a drop of blood falls on leather, many red globules are filtered out from the serum and left on the surface, and, when thus treated, they dissolve, and the coloured solution sinks at once to the bottom of the cell, without coming in contact with the leather. The various spectra may then be observed in the usual manner. This method would be of little or no use if the stain had been wetted, and for a long time I concluded that after such treatment it would be impossible to recognize blood. How- ever, after many experiments, and after having again and again almost given up the inquiry in despair, I found that the difficulty could be overcome in a very simple manner. The best solvent for the insoluble compound of the colouring matter of the blood with tannic acid, is hydrochloric acid diluted with about fifty times its bulk of water. If stronger or weaker, the result is not so good. When a portion of unstained common brown leather is digested in this dilute acid, the solution is scarcely tinged yellow. On adding excess of ammonia, the colour becomes pale purple, or neutral tint, made deeper when the double tartrate and the ferrous salt are added, but remaining nearly clear. This gives a spectrum very dull all over, but without any trace of definite bands in any part. The depth of colour varies much with different specimens of leather. A portion of similar material soaked with wetted blood, gives a yellow solution, made brown-purple and turbid by the Spectrum Method of Detecting Blood. 15 double tartrate and ammonia, and remains so when deoxidized. The band of deoxidized hzematin can however be distinctly seen with a light sufficiently strong to penetrate the turbid and dark solution. Before examining the suspected stain, 1t would be well to make out how much of the unstained leather could be used without giving too dark a solution, and to use no more of the stained. If the deoxi- dized solution be too turbid, the cell may be kept for a while hori- zontal, until the deposit has subsided sufficiently to allow the principal absorption-band to be seen ; but it is not so distinct, when all has subsided, as though the greater part of the hematin still existed as a compound insoluble in dilute ammonia. The presence of tannic acid in wood and other substances might make it necessary to employ a similar process, if the relative amount of blood were so small, that none couid be dissolved out by water, or dilute citric acid. Cases might occur when it would be necessary to decide whether blood were present, along with some other coloured substance, soluble in water. The method to be employed would depend much on the nature of this impurity. If it were a colouring matter, belonging to what I have described in former papers as group A, in which the absorption is removed by sulphite of soda, in an alkaline solution, there would be no difficulty in seeing all the spectra. Thus, for example, it is easy to add so much magenta to the solution of a little blood, that its absorption-bands are entirely hid; but a small quantity of sulphite of soda so completely removes the colour of the magenta, that the various spectra of the blood may be seen almost as well as if it had been pure. The colouring matters of my group B that are most likely to occur, are those of fruits, and in them the presence of the free acid would be almost certain to have changed the hemoglobin into hematin. The best plan would then be to add excess of ammonia, and, if the solution were made too dark, to dilute it with so much water that the strongest light at our command would show the green part of the spectrum sufficiently bright to prove that no absorption-band occurred there. On deoxidizing in the usual manner, the solution may be made somewhat darker by the presence of tannic acid, but the darker band of deoxidized hematin could be recognized without material difficulty. By far the greater number of the colouring matters belonging to my group C are yellow and orange-coloured ; and, since these chiefly absorb the blue rays, they do not interfere with our seeing the bands of the blood spectra, which occur in the green. Cochineal is one that requires special attention. The addition of ammonia to its solution in water gives rise to two bands in the green, which, though differmg materially from those of blood, are yet so nearly in the same situation, that they completely disguise the presence of VOL. VI. Cc 16 Spectrum Method of Detecting Blood. a small amount of blood. However, on adding a small excess of boric acid, the bands of the cochineal are made more faint, and very considerably raised towards the blue end, so as to leave the red end of the green clear, whilst those of oxidized hemoglobin are not changed, and that nearer the red end, if not both, can be seen perfectly well. By proceeding in the usual manner, there is no great difficulty im recognizing the darker band of deoxidized hematin. Other special difficulties might occur in particular instances, but I trust that these examples will suffice to show how they may be overcome. Ido not now know of any that require special remarks ; and, as far as I am able to judge, we need never despair of detecting blood, so long as any hematin remains undecomposed. Fortunately it resists decomposition so well, that this would rarely happen in ordinary circumstances; but yet there are cases in which it does occur, as, for example, when acted upon by strong ozone, or other powerful oxidizing reagents. It is quite possible that stained garments might have been washed, and some of the water employed might be obtained. If no soap had been used, this water could be examined in a long tube of thick glass, ten inches or more in length, and a quarter of an inch in internal diameter, permanently closed at one end with a circular piece of plate glass, and, when filled, covered over at the other with another glass. For examining solutions in such tubes a small pocket spectroscope, such as recently made for me by Mr. Browning, is extremely convenient, and suitable in every respect. 