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considerable number of red globules might be separated, the deposit on the filter should be dashed with a little water, and this solution either examined by itself, or added to the filtered urine. If the depth of colour in the ten-inch tube be so great, that the yellow end of the green part of the spectrum is absorbed, the urine must be somewhat diluted, or examined in a shorter tube. When the depth of colour is about an average, I find that by this means as little as Todocth part of blood can easily be detected in fresh urine, which is equivalent to about one drop in a pint.
V.-On the Cellular Structure of the Red Blood Corpuscle. By JOSEPH G. RICHARDSON, M.D., Microscopist to the Pennsylvania
Hospital, Philadelphia, Pennsylvania. For many years after the magnificent cell theory was first accepted by physiologists, the doctrine of Schwann, who regarded the red blood disks as minute membranous sacs containing a coloured fluid, passed almost unquestioned ; but of late, especially since more careful microscopic observations have become customary, it has been found that the supposed bursting of these little bladders, long looked upon as one of the strongest proofs of their cellular nature, does not take place, and at the present time some of our leading authorities, both in America and in Great Britain, assert positively that the coloured blood disks are non-vesicular, and deny any
differentiation of their substance into cell wall and cell contents.
Thus, for example, Professor Austin Flint, jun., in the second volume of his great work on the ‘Physiology of Man,' p. 116, remarks :—“The structure of the blood corpuscles is very simple. They are perfectly homogeneous, presenting in their normal condition no nuclei or granules, and are not provided with an investing membrane. A great deal has been said by anatomists concerning the latter point, and many are of the opinion that they are cellular in their structure, being composed of a membrane with viscid semifluid contents. Without going fully into a discussion of this point, it may be stated that few have assumed actually to demonstrate this membrane, but they have for the most part inferred its existence from the fact of the swelling, and, as they term it, bursting on the addition of water; and particularly, as it seems to me, to make the blood corpuscles obey the theoretical laws of cell development and nutrition laid down by Schwann. Their great elasticity, the persistence with which they preserve their bi-concave form, and their general appearance, would rather favour the idea that they are homogeneous bodies of a definite shape, than that they have a cell wall with semi-fluid contents; especially as the existence of a membrane has been inferred rather than demonstrated."
Professor Lionel Beale observes, on p. 169 of his work entitled "The Microscope in Practical Medicine: —“The red blood corpuscle of man, and mammalia generally, consists of a mass of soft viscid matter, perhaps of the consistence of treacle, composed of hæmatocrystalline. It is, at least in certain states, soluble in water, but is only dissolved by serum and the fluid part of the blood very slowly. The outer part of this matter is of* firmer consistence than the interior, especially in the older corpuscles. When the latter are placed in water the more soluble matter is dissolved, leaving the harder external portion." Dr. Beale further recounts sundry considerations which, he says, prove conclusively “that the red blood corpuscle is not a cell.”
The distinguished French physiologist, Professor Ch. Robin, supports similar views, asserting, on p. 697 of ' Dictionnaire de Médecine, de Chirurgie, &c.,' The red blood corpuscles are constituted of a homogeneous mass of globulin which is imbibed by or united molecule by molecule to the colouring matter, or hæmatosine, and a certain quantity of fat and saline materials. In mammals, the whole mass is homogeneous, and without any nucleus after the period when the human embryo, for example, attains a length of about an inch; but previous to that the globules, having a magnitude of from 010 to .011 of a millimètre, possess a little round granular nucleus. In all the oviparous vertebrates the globule, whatever its form, encloses a colourless spherical or oval nucleus, insoluble in water and acetic acid, while the red mass is soluble in these menstrua.”
In the course of some researches of my own, however, “On the Detection of Red and White Corpuscles in Blood Stains,” | I have shown, first, that if a few drops of fresh blood be stirred up in many times its bulk of pure water, the coloured-hæmato-crystallin will be dissolved, while a whitish insoluble residue, found under the microscope to be composed of transparent hyaline spheres about 13ooth of an inch in diameter, subsides to the bottom of the vessel; secondly, that if a fragment of dried blood-clot is exposed to the action of a current of fresh water, the hæmato-crystallin will, after a few minutes, be washed away, leaving an aggregation of what appear to be similar delicate cells, altered in shape by mutual
pressure, but still preserving much of their rounded contour; and thirdly, by a calculation of the superficial area of the human red blood disk, based upon accurate measurements of its dimensions when magnified nearly 1800 times, that supposing a cell wall to exist, there would be almost precisely enough membrane contained in it to cover the surface of a sphere having the exact diameter of the red corpuscles when rendered globular by the action of water.
* E. Littré et Ch. Robin, Paris, 1865.
In one of my experiments on the action of water upon blood, as detailed in that paper, the development of Bacteria so obscured these supposed membranous cell walls that they became unrecognizable after standing seventy-two hours, so that, in order to determine whether their apparent insolubility could be overcome by prolonged maceration, I made the following additional investigations :
On the 24th of March, 1870, I thoroughly stirred two fluiddrachms of blood into two fluid-ounces of fresh water, and allowed the mixture to stand undisturbed for forty-eight hours, when a light and flocculent deposit of a pale pink colour, occupying about half a fluid-ounce of the liquid, had fallen to the bottom of the vessel. On examination under the to-inch objective, this was found to be 'chiefly composed of very transparent spherical bodies, about 30th of an inch in diameter, which became beautifully distinct and quite visible with an ordinary 4-inch when tinted by a minute portion of aniline solution introduced at the margin of the cover. In order to prevent the development of Bacteria, about two fluid-drachms of carbolic acid solution were added, and the mixture kept covered in a room of ordinary temperature for four weeks, at the end of which time the delicate colourless spheres were still distinctly visible, although they had a little further diminished in size, only measuring about oth of an inch across.
From these various observations, it appears that human red blood corpuscles are composed of two different ingredients, the one hæmato-crystallin, of a crimson colour, and dissolving freely in water, the other of a whitish hue, and insoluble in water, even on prolonged maceration ; but so minute are the blood disks in mammalia generally, that it is extremely difficult to determine the exact relation of these constituents to each other. It occurred to me, however, that investigations upon the large blood globules of reptiles might be more successful, and after numerous disappointments I procured, in November last, from a former patient near my late residence on Cayuga Lake, in Western New York, two specimens of the Menobranchus or Proteus, whose red blood disks, as far as known, with a single exception, exceed those of all other animals in magnitude, measuring about both of an inch in length by aboth of an inch in breadth, and actually visible, in a strong light, to the naked eye of a myopic person like myself
. The gigantic corpuscles being about six times the diameter, and consequently 216 times the magnitude of those of man, evidently afford much better opportunities for the detection of their membranous parietes, if such exist; and in addition to this great advantage, I discovered, quite unexpectedly, in the course of my experiments upon them,
that their coloured portion possessed the remarkable property of crystallizing with great readiness within its envelope, and so enabling us to analyze, as it were, the corpuscle, by furnishing a singularly positive demonstration of the existence of a cell wall, totally distinct from the cell contents which undergo crystallization. These crystals, as often happens with those produced in the presence of organic matter, are frequently irregular, but their typical form appears to be that of a quadrangular prism, with dihedral summits, the angles sometimes being truncated. They may be easily prepared, as I have now done at least fifty times, by depositing a drop of blood from the Menobranchus upon a slide, allowing it to remain uncovered about ten minutes, or until a mere line of desiccation appears at the margin, and then covering it with a thin glass; on examination with a power of 200 diameters, numerous corpuscles along the edge of the drop where the liquor sanguinis has become most concentrated, will be frequently discovered to contain one, two, or more crystals; and under the most favourable circumstances of temperature and hygroscopic condition of the surrounding air, I have seen this process of crystallization go on until the contents of almost every corpuscle assumes the crystalline form, either wholly or in part, the cell wall being left in the former case perfectly colourless and transparent.
The effect of these crystals as they gradually elongate is very remarkable and interesting, being precisely that which would be produced by sticks of similar shape contained within an ordinary bladder partly filled with fluid ; thus, for example, I have several times seen a single crystal, as if increased in length, thrust out the ends of the oval corpuscle, until the conjugate diameter of the cell became one-third greater, while its transverse dimension diminished to less than half its original magnitude, the nucleus being compressed closely against the side of the prism. Or in cases where one or more crystals happened to lie across the long axis, that decreased until the whole corpuscle assumed a lozenge-shaped or rectangular form, as in a very perfect specimen which I have mounted dry, the folded edge of whose capsular membrane may be seen supported by the crystals, like a washerwoman's clothes-line upon its prop. It
may in the first place be objected to this demonstration, that the appearance which it affords of a plicated membrane around the extremities of the crystals is caused by partial desiccation of the surface of the corpuscle while the specimen was being prepared; that such cannot, however, be the case, is proved by the fact that if, to blood freshly drawn from the reptile upon a slide, water is added, beneath the microscope we can produce an exosmosis of the coloured material into the diluted liquor sanguinis, leaving the same transparent cell wall, which becomes visible when the cell contents are crystallized within it; and it is obvious that a membranous envelope, which is equally distinct under the opposite states of dryness and moisture, cannot be considered the result of either condition. Again, perhaps it will be asserted, secondly, that the appearances here presented might be simply the result of partial crystallization in such a drop of viscid material as Professors Flint and Beale consider the red blood disks, which drop, if the process were complete, would have entirely assumed the crystalline form; but I think I can quite destroy the force of that or any similar argument by the aid of other mounted preparations, some of them showing that well-developed crystals, which happen to lie in favourable positions, may include almost all the coloured portion of the corpuscle, without in the least affecting the contour of its cell wall.
I regret exceedingly that the difficulty of obtaining and preserving the Menobranchus alive, has prevented me from attempting to exhibit specimens of its fresh blood; but in the hope that other microscopists will repeat and correct or confirm my researches upon it, I am desirous of recording them and the conclusion which they seem to involve.
After a great many attempts, on which I spent altogether about eight hours steady work, I have twice succeeded in cutting a corpuscle in two with sharpened needles upon a stage of the microscope, and beneath a half-inch objective, combined with a No. 2 eye-piece. On penetrating the vesicle with the edge of the needle, its coloured contents were instantly evacuated, and disappeared at once in the surrounding fluid, while the cell wall immediately shrunk together, and became twisted upon itself, and around the nucleus into a perfectly hyaline particle, which showed some tendency to adhere to the point of the instrument. It would therefore seem that the hæmato-crystallin was neither viscid nor semisolid, and that the cell wall was structureless, and possessed only moderate tenacity, but of course the observations were too few in number to be accepted as conclusive.
When the corpuscles remained for two or three hours under observation, those which did not crystallize, often showed the wrinkled appearance figured by Hassal in his Illustrations, and described by Rollett, in Stricker's Handbuch der Lehre von den Geweben,' Zweite Liefrung S. 286, and which seemed to me due to the tendency of their colourless envelope, as the contained hæmatocrystallin condensed around the nucleus, to accommodate itself to the diminished contents of the cell by falling into folds frequently ramifying from the nuclear centre. When pressure was made by means of a mounted needle upon the covering glass, almost directly over a red disk, whose contents had undergone this contraction, the first effect was to round out the contour of the corpuscle, and