USA > Ohio > Hancock County > Findlay > Twentieth Century History of Findlay and Hancock County, Ohio, and Representative Citizens > Part 135
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I.
1. To a large quantity of sputum add antiformin to a 25 per cent solution.
2. After the production of a homogeneous solution add 96 per cent alcohol, thereby reducing the specific gravity and hastening sedimentation.
3. Pour off supernatant fluid.
4. Wash sediment in centrifuge.
5. Smear and examine.
II.
1. Take equal parts of sputum and sterile water.
2. Add antiformin to 2 per cent solution.
3. Allow to digest and sediment to accumulate.
4. Pour off supernatant fluid and smear directly.
In II washing is not necessary owing to the low percentage of antiformin; on the other hand, this process requires about three days for completion.
Meyer * also proposes two methods practically identical with the preceding :
* Hüne substituted a mixture of sodium hypochloride 11.1% and potassium hydroxide 5.6% for antiformin with satisfactory results.
.
I.
1. To the sputum add antiformin to a 20 per cent siz
2. Transfer the entire resulting solution to centrifuge and centrifugalize violently.
3. Wash sediment twice in centrifuge.
4. Smear and examine.
II.
1. To the sputum add antiformin to 3 per cent solo: : examine sediment without washing.
Seeman " uses either the centrifuge method or gravity -. mentation. He also does away with the necessity of va the sediment by using either diluted egg albumin or x- some of the original sputum to aid in fixing the smear.
METHOD.
1. To the sputum add antiformin to a 15-20 per cent .. tion.
2. Add 96 per cent alcohol after digestion is complete.
3. Smear sediment directly, using either egg albuni: some of the original sputum ; or
2. Centrifuge the material.
3. Smear as in 3.
In using some of the original sputum to aid in firing : smear, Seeman not only does away with the necessity of w- ing the sediment, but at the same time gains some ins .: into the general composition of the sputum, as regards : number and character of the other organisms, the type if- predominating, and the presence or absence of elastic flv and also renders the search for the tubercle bacillus a: easier by having the blue field upon which to focus.
Thilenius' uses the centrifuge after first having added i hol to the sputum-antiformin mixture. He also uses .. formin in from 10-50 per cent solution.
Haserodt' recommends the use of ligroin, a light by" carbon oil, for obtaining the tubercle bacilli. His meth: as follows :
1. To the sputum add antiformin to make 5 per cent 9. tion.
2. Add a few cubic centimeters of ligroin and shake til emulsion is produced.
3. Allow to stand until a sharp line of separation (cc. between the ligroin and the sputum-antiformin mixture.
4. With platinum needle remove several loopsful of mater. from just below the ligroin; spread this on a slide and ri- amine in the usual manner.
The tubercle bacilli and other undissolved particles in . mixture are carried upward by the little droplets of lig. and deposited just below the line of separation between th- two solutions. Haserodt claims that the smears, by 2. method, show no tendency to wash off the slides, but this si- ment is not upheld by other investigators. The method no advantage over the previous ones, and has failed to ff as good results.
Bernhardt's " procedure is exactly the same as Haservit's except he uses 5 cc. of sputum and adds 20 cc. of a ?i . cent solution of antiformin.
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.. Equal parts of sputum and water.
!. Add antiformin to a 25 per cent solution, shake vio- tly.
3. After 4-6 hours gently turn vessel to dislodge any par- es which may be adhering to the side of the vessel.
1. Allow to settle for several hours.
i. Pour off supernatant fluid.
i. Fix sediment to slide with egg albumin or some of the ginal sputum.
Che conclusions, finally reached from a review of the sev- I articles dealing with the use of antiformin in sputum minations, may be briefly summarized as follows:
. Large quantities of sputum should be used.
. Sputum should be diluted with sterile water.
. Antiformin should be used in from 15-25 per cent itions.
. The sputum is dissolved in from 10-45 minutes-the illi, however, are uninjured after days of exposure in these itions.
. The resulting solutions should be watery in consistency either colorless or pale yellow-if need be, more water or 'e antiformin may be added.
. The tubercle bacilli may be obtained from the solution er by gravity sedimentation, by centrifugalization, or by use of ligroin. Sedimentation may be facilitated by the ition of alcohol to lower the specific gravity. Of these :ral methods, gravity sedimentation is perhaps most used.
. Washing of the sediment is unnecessary if egg albumin, ited with 10 parts of water, plus 1 per cent formaldehyde; some of the original sputum be used to fix the smear.
. The smear stains perfectly by the Ziehl-Neelsen method.
. In examining the smears one may find acid-fast rods ch are to be differentiated from the tubercle bacillus by g larger, and straighter, and especially by the fact that · readily transmit light. These rods are probably fat lles, as suggested by Goerres.
). In the entire smear there may be only two or three lli-but these are sufficient for a positive diagnosis.
L. Finally, by the use of antiformin in the examinations putum for tubercle bacilli one is enabled to demonstrate 1 in from 7.3 to 17.2 per cent of the specimens reported gative after careful smear examination by the old method. may be seen in the following reports:
eyer (loc. cit.) in examining 113 sputa, found 14 posi- by the ordinary smear examination. Of the remaining o-called negative specimens, 14 or 14.1 per cent were onstrated to contain tubercle bacilli by the antiformin .od.
aserodt (loc. cit.), in 340 examinations, classed 300 as tive after careful examination by the ordinary method. le antiformin method 22 or 7.3 per cent of these negative , were shown to contain tubercle bacilli.
erres (loc. cit.), in 296 examinations by the ordinary od, found 162 negative; 28 or 17.2 per cent of these were itely positive by the antiformin method.
Some months ago the writer undertook a study of the anti- formin method of sputum examination to determine its value and limitations.
After trying various procedures, the following was finally adopted as the most satisfactory both as regards method and results.
1. Place the entire 24 hours' sputum in a conical settling glass; if the amount be excessive it is perhaps better to use only 15-20 cc.
2. If the specimen is thick, add an equal volume of dis- tilled water. Less tenacious specimens do not require so much dilution .*
3. Add an amount of antiformin equal to one-fourth the volume of the diluted sputum; in other words, sufficient to make a 20 per cent solution.
4. Stir thoroughly, thereby breaking up the masses of mucus and greatly hastening complete solution.
5. Allow to stand till solution appears homogeneous. It should now be watery in consistency and pale yellow in color ; if necessary, more water or more antiformin should be added and digestion allowed to continue. This will usually require from a few minutes to an hour, but may be allowed to con- tinue for days with no resulting harm to the tubercle bacilli.
6. Add an equal volume of 95 per cent alcohol. By this procedure the specific gravity is reduced from about 1.030 to below 1.000; thereby not only hastening sedimentation, but making it more complete.
7. After stirring, allow to stand till sedimentation is com- plete. This will occur in 2-4 hours, but a period of 12-24 hours is recommended. During this sedimentation it may be necessary to gently turn the vessel to dislodge little particles of sediment which may be adhering to the sides of the vessel. 8. Pour off this clear supernatant fluid.
9. Make smear from the sediment on a glass slide, using some of the original sputum to aid in fixing the smear. This is best done by making a smear from the sputum before anti- formin is added and afterwards spreading the sediment from the sputum-antiformin mixture on the same slide. Stain and examine in the usual way.
If one has the proper laboratory facilities the centrifuge may be used to hasten the work, or to remove the last trace of doubt as to the accuracy of the results; but from the reports of the other observers and my own exeprience, if tubercle bacilli are present, they will appear in the sediment obtained after adding alcohol. The use of the centrifuge merely shortens the time element, but in so doing adds much to the difficulties of the method.
Antiformin has been used in the examination of 103 sputa. Twenty specimens were obtained from the patients at Eudo- wood Sanitarium through the courtesy of Dr. Sloan, three from the Epstein Hospital, through the courtesy of Dr. Grover, and the remaining from the patients in attendance at the Phipps Dispensary of the Johns Hopkins Hospital. For the
* Here it might be well to make sure that the distilled water harbors no acid-fast organisms.
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collection of the sputum wide-mouthed bottles of 100 cc. capacity were used, these being carefully washed in distilled water before being distributed. The patients were instructed to expectorate directly into the bottles, beginning with the first morning expectoration and continuing to use the bottles for the next 24 hours, or until they had accumulated from 15- 20 cc. of sputum. However be it here noted that in this re- gard the results were very disappointing, as the dispensary patients frequently returned the bottles with but two or three cubic centimeters of sputum and in many cases this was with- out doubt entirely from the upper air passages. The results from the sanitaria were more satisfactory.
Dividing the cases examined according to the clinical diag- nosis, 73 were definite pulmonary tuberculosis, 6 were probable pulmonary tuberculosis, 16 were doubtful pulmonary tuber- culosis and 8 were definitely non-tuberculous.
By the old method all the sputa of the non-tuberculous, doubtful and probable groups, 30 in number, were negative. Of the 73 clinically positive cases, 31 showed tubercle bacilli in the smears examined.
By the antiformin method the examination of the same specimens showed no tubercle bacilli in any of the non-tuber- culous, doubtful or probable cases. In the 31 cases positive by the old method great enriching was seen. In the ordinary smear an average of 25 bacilli could be counted in two minutes. In the antiformin smear an average of 138 tubercle bacilli could be counted in the same space of time. Of the 42 re- maining specimens from clinically positive cases which, by ordinary smear examination, were negative for the tubercle bacillus, seven or 16.6 per cent were demonstrated to contain
the bacillus by the antiformin method; or considering al : positive, probable and doubtful cases, a total of 95; 64 my negative by the old method, but 10.9 per cent of these ral demonstrated to contain tubercle bacilli by the antifez; method.
I therefore feel fully justified in heartily recommend- the use of antiformin in the examination of sputum : tubercle bacilli, not only in hospitals and dispensaries, : in the laboratory of every private physician. The gears adoption of this method, combined with repeated eranie tions, will do much towards reducing the percentage of (už clinically diagnosed as definite pulmonary tuberculosis, E which however an absolute diagnosis cannot be made. 01 5: greater value than the finding of tubercle bacilli in the spr. of cases clinically diagnosed as definite pulmonary tobe. losis, is the possibility which this method offers of den. : strating bacilli in the cases still classed as only probable : doubtful, thereby enabling us to enforce a vigorous and eis- matic treatment at a time when such treatment is of 2.5 avail.
LITERATURE.
1. Berl. klin. Wchnschr., 1886, XXIII, 713-717.
2. Ztschr. f. klin. Med., 1910, LXX, 86-101.
3. Berl. klin. Wchnschr., 1908, XLV, 1346-1349.
4. Deutsche med. Wchnschr., 1909, XXXV, 1791-1793.
5. Tuberculosis, 1909, VIII, 71-75.
6. Berl. klin. Wchnschr., 1909, XLVI, 628.
7. Berl. klin. Wchnschr., 1909, XLVI, 615; 1169.
8. Hyg. Rundschau, 1909, No. 12.
9. Deutsche med. Wchnschr., 1909, XXXV, 1428-1429.
SERUM TREATMENT OF HEMORRHAGIC DISEASES.
By W. L. Moss, M. D., Associate in Medicine, Johns Hopkins University,
AND J. GELIEN, M. D.
(From the Research Laboratory, The Phipps Tuberculosis Dispensary, The Johns Hopkins Hospital.)
Under the designation "Hemorrhagic Diseases " is grouped a number of widely different morbid conditions, having hemorrhage as a common feature. The bleeding varies in location and in severity from insignificant cutaneous hemor- rhages in mild cases of purpura, to appalling and even fatal losses of blood in more serious conditions.
An enumeration of the diseases with which hemorrhage is, or may be associated and which are sometimes designated as hemorrhagic diseases includes :
Hemophilia, hereditary and spontaneous.
Hemorrhagic diseases of the new-born, of which there are doubtless several forms.
Purpuras, acute, chronic, simple, rheumatic, senile, etc. Jaundice. Grave anemias.
A heterogeneous group, including nephritis, typhoid, 2%. other severe infections.
Is hemorrhage in these conditions dependent upon a (iz- mon cause or are different factors operative? At present r- are unable satisfactorily to classify hemorrhagic disease we do not know the causes of the hemorrhagic diathesis. : in order to study this subject intelligently, a consideration ! the theories of the coagulation of normal blood is necessary There are almost as many theories concerning coagulation es there are investigators in this field. If we content oursele with stating the points upon which all are agreed our task easy. The coagulation of the blood depends upon the acti of fibrin ferment, which normally is formed only after } blood is shed, on fibrinogen, which exists as such in the cur- culating blood. This much is fairly simple. Fibrinogen is :
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ad its characteristics are fairly well determined. The part plays in coagulation is known, but since hemorrhage is a ithological condition and coagulation does not take place der normal conditions, this action of fibrinogen can scarcely : taken as representing a normal function. Concerning its igin, the use to which it is put under normal conditions, and s fate in the body, we know little with certainty. The origin ad mode of action of fibrin ferment is enveloped in still reater obscurity, although something is known of the finished roduct itself.
The term fibrin ferment is now used less than formerly, as le ferment nature of this body has been seriously called into lestion. Thrombin, as it was designated by A. Schmidt, has en isolated in approximately pure form and its properties udied by Howell.' Thrombin added to a solution of fibrino- In, causes coagulation by the formation of fibrin. Thus we ive as factors in the much studied coagulation of the blood ro substances, fibrinogen and thrombin which together form ›rin, the essential feature of the blood clot. Not only is the igin of thrombin the subject of many conflicting theories, it also the mode of action of thrombin on fibrinogen in the oduction of fibrin. Whether it is a ferment, a chemical, or physico-chemical action is still undetermined.
Thrombin does not exist as such in the circulating blood, it since normally it appears within a few minutes after the ood is shed, it seems evident that all of the constituents nich enter into its formation must be present in the blood fore it is shed. Most theories concerning coagulation postu- te, therefore, as a normal constituent of the blood, a mother bstance of thrombin, usually designated prothrombin or rombogen. While there can scarcely be a doubt as to the istence of prothrombin, it has never been identified or iso- ed. To account for the conversion of prothrombin into the tive substance, thrombin, the presence of a ferment has en assumed, and to this ferment Morawitz gives the name rombokinase. Since it has been found that blood does not. igulate if the calcium which it contains is precipitated in oluble form as soon as the blood is shed, soluble calcium ts have been recognized as essential to the action of throm- cinase in converting prothrombin into thrombin. This lase has never been isolated and its existence is really a mat- of theory. Calcium salts of course do exist in the blood 1 their presence can be quantitatively determined, but the ct part they play in coagulation remains a subject of con- versy.
)ther substances which have been postulated in various ories to account for the coagulation of the blood, such as ombozym, zymoplastic substances, etc., need not be con- red here, but it is necessary, for the purpose of this paper, jo one step further into the theories concerning the forma- L of thrombin. We may accept as a necessary part of any ›ry, the existence of prothrombin normally in the circulat- blood. Something happens when the blood is shed which ses the prothrombin to pass over into thrombin and since have no more satisfactory explanation of this transforma-
ject. There is one change in the blood which we know begins to take place as soon as it is shed, and this is the disintegra- tion of the platelets, and less rapidly, the disintegration of the leucocytes. It is not unnatural, therefore, that investigators have sought the origin of thrombokinase in these elements.
We have attempted to develop from the literature, a more or less general and somewhat composite theory of coagulation. It serves to illustrate the highly speculative character of much that is held concerning this important phenomenon and indi- cates the difficulties to be overcome in arriving at the etiology and classification of the hemorrhagic diseases.
Hemophilia is the most striking and most interesting disease in this group and it is natural that it should have been most studied. To account for the hemorrhages many theories have been advanced. These vary from such simple ex- planations as abnormal thinness of the blood vessel walls, lead- ing to rupture under slight provocation ; abnormally high blood pressure; a disproportion between the total amount of blood and the total capacity of the vascular system; to such pro- found and obscure explanations as that "in hemophilia we have to do with an inherited chemical (fermentative) degen- eration of the protoplasm of the formed elements of the blood, and perhaps of the whole organism " (Morawitz and Lossen ').
It is not necessary to enter into a discussion of the various explanations which have been offered for the hemorrhage in hemophilia and other hemorrhagic diseases. Some have thought that there is a lack of one or more of the factors nor- mally present and concerned in bringing about coagulation ; others have considered that there are present in the blood of these patients abnormal substances which are antagonistic to coagulation. Thus a lack of fibrinogen, of available calcium salts, of prothrombin, of thrombokinase, or of the elements from which thrombokinase is supposed to arise, on the one hand, and antithrombin and other anticoagulants on the other hand, have been held responsible for the failure of the blood to coagulate. Thus it may be seen that until more exact in- formation is obtained as to the cause of the hemorrhagic diathesis any measure directed towards its treatment other than mechanical must be largely empirical.
When one comes to consider hemostatic measures one finds a long list for disposal. Excluding the direct measures, liga- tion, tamponading, etc., which are applicable to accessible hemorrhages in normal individuals, less successfully often- times in those with a hemorrhagic diathesis, we have the fol- lowing : administration of calcium salts to increase the coagu- lability of the blood; injection of gelatin to increase its vis- cosity ; administration of adrenalin to narrow the vessels. Un- fortunately, the latter drug raises blood pressure, unless applied merely locally and while the local application does sometimes arrest hemorrhage, its use is accompanied by the. danger, that if the hemorrhage does not cease entirely during the time the vessels are constricted, it is apt to become more profuse upon the subsequent dilatation, which occurs after the constricting action passes off.
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Ergot probably has little value as a hemostatic aside from its use in obstetrics.
Other drugs which are classed as hemostatic are ferric chloride, silver nitrate, hydrastin, hydrastinine, gallic acid, lead acetate, etc. None of these, however, have proven very successful in the conditions under discussion.
Recently the treatment of hemorrhagic diseases has at- tracted renewed attention. It is to the investigations of Weil that the present trend of the work is usually ascribed. In 1905 he published his investigations concerning the patho- genesis and serotherapy of hemophilia,3 which have been fol- lowed by numerous papers by himself and others.4, 5, 6, 7, 8, 9
Weil's work may be very briefly summarized as follows: The hereditary and spontaneous forms of hemophilia can be differentiated in origin, severity of clinical manifestations, character of the blood and its mode of coagulation. (The faulty coagulation in the hereditary form is referred to the presence in the blood of anticoagulants; in the sporadic form, to the absence of substances normally present and essential to coagulation. He considers the changes of the blood in marked hemorrhagic conditions as analogous to, but not identical with, those of hemophilia and recommends, as a cure for the sporadic type of hemophilia and for many of the other hemorrhagic states and as the best therapeutic agent available in the treatment of hereditary hemophilia, subcutaneous in- jections of 30 cc. or intravenous injections of 15 cc. fresh human or animal serum. Elicagaray 10 confirms Weil's work.
Baum 11 repeated much of Weil's experimental work, most of which he was unable to confirm. In reviewing his ex- perience he states that he was much encouraged by his first case in which local applications of serum apparently stopped the hemorrhage, following the extraction of a tooth, in a hemophilic. In his succeeding cases, however, and in his animal experiments local applications of serum seemed en- tirely without effect. As to injections he found that in hiru- dinized rabbits, the coagulation time was much reduced by in- jections of serum, except in cases in which the blood had been rendered incoagulable to a marked degree by hirudin.
Baum treated three cases of hereditary hemophilia by means of serum injections with negative results, but thinks that in other hemorrhagic conditions good results may follow its ad- ministration.
Trembur 12 reports one case of hemophilia successfully treated by injections of sheep and rabbit serum.
Lommel18 reports the successful treatment of hemorrhages in a four-year old hemophilic by the use of subcutaneous in- jections and local applications of antistreptococcic serum.
Class 14 reports one case of purpura hemorrhagica, which he considered cured by injections of antidiphtheritic serum.
Leary 15 reports a series of twenty cases treated by means of serum injections. His cases may be summarized as fol- lows :
1. Jaundice, nine cases. The first case did not receive serum until after the hemorrhage had set in, following an operation. This patient died. Each of the remaining eight cases received before operation, as a prophylactic measure, 30
cc. rabbit serum subcutaneously. All but one of these : covered with but little bleeding following the operation : the fatal case the author ascribes the death not to hemorras but to cholemia.
2. Hemorrhage of the new-born, three cases, two recoveres one death.
3. Hemophilia, one case received diphtheria antitoxin : bit serum, and finally direct transfusion of human bis Patient regained health rapidly.
4. Purpura, one case, cured.
5. Post operative hemorrhage, two cases, one cured. : died.
6. Uterine hemorrhage, one case, cured.
7. Typhoid fever hemorrhage, two cases, cured.
8. Ruptured tubal pregnancy (operated) one case, cured. Among others who speak favorably of the use of serum : jections in the treatment of hemorrhagic diseases is Wel who reports twelve cases of hemorrhage in the new-born : of which he claims were cured by subcutaneous injection: fresh human serum. He injected as much as 209 cc. ix : days, but averaged about 80 cc., in 10 cc. doses, exteriz. over four days.
Schockaert 17 used subcutaneous injections of antistrer- coccic serum successfully in two cases. One was a very aniz patient who bled profusely under operation. Fifty cubic a: timeters of serum were injected and the author reports the immediately the hemorrhage stopped, so that the operat .: could be concluded with a dry field of operation. The sarz. case was one of rebellious menorrhagia which resisted hydro- tis, ergot, stipticine, etc., but was controlled by injections : 10 cc. of antistreptococcic serum at the beginning of the De. strual periods.
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