Complete Text (Part 1)

Class Book Rf^V? i.^ ]0 GjpiglitlN" CDEffilGHT DEPOSm MANUAL OF LABORATORY DIAGNOSIS Compiled and Elaborated by Herman John Bollinger, S. B., M. D. Assistant in Bacteriology Johns Hopkins University Preface by Sidney R. Miller, S. B., M. D. Associate in Clinical Medicine Johns Hopkins University BALTIMORE. MD. MEDICAL STANDARD BOOK CO, .i'^ ^ iP^\ Copyright 1919 Medical Standard Book Co. Baltimore, Md. mR "5 19 Press of , > ' The Medical Standard Book Co. Baltimore CI.A5L2476 This book is compiled irom lectures given by the following instructors at the Johns Hopkins Medical School. S. R. Miller, M. D., Lectures on Urine, and Blood. Marjorie D. Batchelor, M. D., Lectures on Stomach Analysis. F. A. Evans. M. D., Lectures on Sputum and Stools. W. A. Baetjer, M. D., and S. R. Miller, M. D., Lectures on Blood. C. G. Guthrie, M. D., Lectures on Parasites. PREFACE. Requests have come from time to time for out- lines of the general course in Clinical Microscopy, as given at the Johns Hopkins Medical School, or for summaries of the individual subjects covered. Such outlines have not any particular merit other than that they serve as guides for teaching, or compends, useful for the quick reviewing of any particular topic. It has always seemed a wiser policy to en- courage each student, carefully to take his own notes and prepare for himself such a working out- line as best suited his own needs. Condensed notes and compends in general are too prone to stimulate superficiality to warrant their unqualified recom- njendation. Moreover the practice here has been to have each instructor cover a different subject each year, thereby ^\idening the scope of each man's knowledge and interests; each improving on his predecessor's lectures, if possible. Consequently, no set plan is followed and stereotyped lectures have been consistently avoided. The present summary of the lectures given in the past year, is published chiefly as a result of the en- thusiasm and painstaking work of one of the stu- dents. It is not intended that these notes shall be comprehensive: the interpretation of tests, etc., has not been given much space, and, by reason of the universal "war-time lack of time," careful correction has not been possible. The notes are presented merely as representing the essential subject m^^tter covered in the lectures on urine analysis, exaniina tion of the gastric contents, sputum, feces, parasi- tology and hematology. The publication of these notes had the sanction of the late Dr. Theo. Janeway, whose interest in the application of clinical labora- tory methods to the diagnosis of disease was always keen and enthusiastic. It had been his intention to write this preface as evidence of his approval of the project. B}' reason of their absence in France, Drs. W. A. Baetjer, C. G. Guthrie and Frank Evans have been unable to review the sections previously cov- ered by them. Knowing full well their great interest in the subject, it has seemed wise for me to stand sponsor for them all, and to permit publication, but only with a clear understanding of the limits of the book, the constant changing of methods and inter- pretations, and the lack of balance unavoidable in a work of this kind. Sydney K. Miller^ M.D. The Clinical Laboratory, Johns Hopkins Medical School. ALBUMINURIA Definition: The accumulation of a coagulable protein in the urine. The protein has, in true cases, escaped through the renal parenchyma. Two groups : 1. True condition in which the renal parenchyma is faulty. 2. False. Condition in Avhich the protein has been added to the urine during its passage through the G. U. tract. Origin : 1. Blood protein. 2. Secretion or degeneration of renal cells. 3. Substances added along G. U. tract. No chemical method has been devised to differ- entiate these ditlerent forms. Most of protein is of blood origin all agree. Serum albumin. Greatest in frequency and amount. 5 to 40 grams per day. 20 to 100 mg. oc- curs normally, but is not detectable. Albumin is only clinically present when it can be detected by the ordinary laboratory means. This is set as a standard. Serum albumin is soluble in water, co- agulable by heat, precipitated by alcohol and salts of the heavy metals, and by saturation with magnesium sulphate, and is Levo-rotatory 62 degrees. Serum globulin. This is always present with serum albumin. It represents from 10 to 75 per cent of total protein. It is insoluble in water and dilute acetic acid in the cold. It is precipitated by saturation with magnesium sulphate and half saturation with am- monium sulphate and partially precipitated by so- dium chloride saturation. Pseudo-glohuliUy euglo'dulin^ and fibrinogen. These belong to the globulin group. Fibrinogen is practi- 8 cally never en countered and when it is is perfectly obvious, for the urine undergoes spontaneous clot- ting. Euglobulin and pseudo-globulin are probably the same as nucleo-albumin. They are of no practi- cal importance. jSiucleo-histones are of no practical significance. Bence-Jones protein is protein sui generis. Alhumoscs are characterized by a tendency to be soluble upon heating and insoluble upon cooling. They are increased in acute types of nephritis and in the absorption of exudates. OCCURRENCE OF ALBUMINURIA. 1. With renal lesion. 2. Without renal lesion. Occurrence of albuminuria without definite renal lesion. 1. Severe muscular exercise. 2. Severe emotional upsets. 3. After cold baths. 4. Alimentary ingestion of excessive amounts of proteins. 5. New-born, first 8 to 10 days of life. 6. Pregnancy, toward the end or after parturiton. 7. Adolescence, 12 to 16 years of age. 8. Hypostatic, associated with large spleens, no- ticeable after the patient has lain down for some time. 9. Cyclic, orthostatic, or lordotic. This type oc- curs in young adults, 12 to 20 years of age. Albu- min appears only after the patient has been up and about for some time. Early morning urine is al- bumin free. These patients usually have abnormal lordosis, and w^hen corrected the albuminuria clears up. According to Jehle there is a deficiency of renal function while the patient is standing, which clears up when he lies down. The condition does not predispose one to nephritis, bnt tends to clear up as the x^atient grows older. Salient features of functional disturbance. 1. Albumin slight. 2. Occurs chiefly in young people. 8. Transient or intermittent. 4. Occurs without other evidences of renal disease. 5. No renal disease etiology. 6. Patients tend to be a bit peaked and have a neurotic tendency, with vasomotor instability, der- matographia, etc. Alhurninuria in those with transient renal injury. 1. Fevers (with casts). Disappears with the dis- appearance of the fever and patient does not suffer inconvenience during rest of life. 2. Toxic. Caused by drugs and secondary to other states such as pj'^orrhea, genito-urinar^ infec- tion, etc. Dental treatment, etc., may clear up con- dition. 3. Hematogenous. Caused by altered condition of blood as in leukaemia, pernicious anaemia, pur- pura, scurvy, beriberi, and jaundice. 4. Traumatic. Caused by injury in the vicinity of the kidney or in remote parts of the body. Occurs in epilepsy, brain tumors, and sometimes after bimanual palpation of the kidney as well as other abdominal manipulations. 5. Intermittent, characterized b}' periodic occur- rence, indicative of: (a) Insidious development of nephritis. (b) Stasis. Summary. Kegard faint traces of albumin in people under 20 years of age in good condition as functional. Ex- 10 amine last voiding at night and iirst in tlie morning to determine whether it is orthostatic, or hypostatic. Use care to exclude extra-renal sources of albumin, bladder infection, vaginal discharge, etc. Regard albuminuria in febrile and toxic conditions as ne- phritis, at that time at least. One cannot judge the extent or the type of nephritis by the frequency of albuminuria. The absence of albumin and casts does not exclude nephritis. Detection of proteins in the urine: Serum albu- min, serum globulin, nucleo-albumin, and albumoses. 1)1 order to detect these substances the iirine must he: 1. Absolutely clear, especially if small traces are to be detected. Make urine clear by filtration or by the us^e of kieselgMhr or animal charcoal. When the urine comes tiirough the filter paper turbid add about 1/10 volume of kieselguhr and refilter. 2. Urine should be faintly acid. ^. Urine should not be too concentrated or too dilute. In cases where it is very dilute 1005 to 1006 specific gravity, it should be made more concen- trated by the addition of about 1/10 volume of so- dium chloride. 4. In doubtful cases don't rely on one test alone. Heat and acetic acid test. After the urine has been made clear by the above methods take a test tube about 2/3 full of the clear urine and heat the upper third in a bunsen flame. If no cloud appears the urine is x)robably normal. If a marked cloud appears it can be : 1. Calcium phosphate. 2. Calcium carbonate. 3. Albumin. In order to differentiate these add 5 per cent acetic 11 acid drop b}' drop. If cloud is due to calcium phos- phate it disappears ; if it is due to calcium carbonate it disappears with the evolution of gas ; if it is due to albumin it persists, Ix-comes greater or becomes slightly less, in accordance Avith the amount of the former substances present. It must be remembered that these protein substances are soluble in an ex- cess of acid or alkali. To detect very faint traces use a dark background. Sources of error: (a) Technique, (b) Other pre- cipitable substances. 1. ^ucleo-albumin. This is precipitated with 5 per cent acetic in the cold. Albumin is only pre- cipitated after heating. 2. Eesinous acids such as cubebs, guiac, etc., give a precipitate with heat and acetic acid. These if shaken with alcohol or ether are dissolved. 3. Albumoses. The precipitate of these comes down after cooling, and reheating gives re-solution. Heat and nitric acid test. Heat urine to boiling and add 20 per cent HNO3 drop by drop. A cloud is fairly indicative of albu- min. It has the same sources of error as the heat and acetic acid test. These two tests are sensitive to .005 grams in 100 cc. Heller's test. Urine and concentrated H]S^Oo are layered in equaJ parts, the urine above and the acid below. A cloud at the line exactly can be : 1. If white, albumin. Its breadth depends upon the quantity of albumin present. xA^fter a time it tends to diffuse upwards. 2. Thymol. This has a grayish or brownish color. There is also a zone of oreenish color below and a 12 reddish one abo\e llic^ line of contnct. ronfirm b}' t^haking with petroleum ether. 3. Albumoses. Tlie cloud given by these is ideii- lical with that of albumin. J>y heating this cioud disappears. 4. Urea nitrate. This cloud is yollowisli brown and is made up of crystals which give a shimnier. A cloud above the line of contact may be: i. A broad, yellowish band about II/2 cm. above line of contact due to urates in concentrated urines. They are dissolved by heating. 2. Xucleo-albumin gives a cloud ^4 to % cm. above the line of contact. It tends to disappear by dilu- tion. By diluting the urine to 1005 to 1008 nucleo- albumin and albumoses can be eliminated. This test is sensitive from .002 to .007 grams per 100 cc. Potassium ferrocyanide and acetic acid test. From 5 to 10 cc. of urine in a test tube to which is added acetic acid. Do not heat. Filter and to the nitrate add 10 per cent potassium ferrocyanide, drop by dro]). Albumin gives a whitish precipitate. Sources of error: 1 . Iron present in large amounts. Color different. 2. Boiling urine in a glass container. 3. Reagent itself. 4. Albumoses which upon heating disappear, 5. ^ucleo-albumin same as albumoses. QUANTITATIVE TESTS FOR ALBUMIN. The most common and easily performed is the em- ployment of the Esbach tube. It differs in accuracy from the Kjeldahl metliod by .3 gm. Urine is added to the mark U and reagent to the mark E. The tube is inverted ten or twelve times to insure thor- ough mixins,- of the contents and is then allowed to 13 stand for about 24 hours and read. The reading below gives the number of grains per liter. Reagents useds 1. Picric acid. This is merely mentioned to be condemned. It not only precipitates albumin but also albumoses, uric acid, creatinin, resinous acids, etc. 2. Tsucliiya'S reagent. Phosphotungstic acid 1.5 grams Concentrated HCl 5.0 cc. 95 per cent alcohol g. s. ad. . . .100.0 cc. The advantage of this reagent is that it eliminates the above error. The disadvantages of the test are that it is inac- curate in urines containing over 4 per cent albumin ; and in some cases the precipitate sticks to the sides, may float or may settle unevenly. OLOBTTLINS: Purdy's test. Globulins are soluble in salt solution, but insolu- ble in distilled water. Dilute the urine and get a cloud, or, better still, drop the urine in a test tube half full of distilled water and watch for a cloud against a black background. Ammomum suliyliata method. Make the urine alkaline with amrnroniura hyd^65^ ide and filter. Layer equal amounts of the filtrate and saturated ammonium sulphate. A cloud at the line of contact is fairty indicative of globulin. For more accurate determination do following test : Urine plus equal volume of saturated ammonium sulphate solution. Let stand an hour and filter. Wash pre- cipitate with half saturated amonium sulj^hate until filtrate is albumin free. To further differentiate it dissolve precipitate in water and heat on Avater 14 bath to coagulate the proteins. Filter and wash the precipitate with water. To the precipitate add 1 per cent solution of sodium carbonate and heat on the water bath. Filter and neutralize with acetic acid. A precipitate signifies globulin (Web- ster). ' - BENCE-JONES PROTEIN. (S. K. Miller and Walter A. Baetjer, J. A. M. A., 1-19-18, vol. 70, pg. 137-139.) Tliis is the most infrequent type of protein found in the urine. In 1816 Bence-Jones described the characteristics of the protein, which were tlie forma- tion of a gelatinous precipitate at a low temperature wliich disappeared upon boiling and reappeared uj)on cooling. In 1876 Rustizky described multiple myeloma. In 1889 Kahler described a case of mul- tiple myeloma with Bence-Jones protein in the urine. The association of multiple myeloma with Bence-Jones protein is almost constant but not spe- cific, for it does occur in other conditions. In no case, however, where Bence-Jones protein has been found has the bone-marrow been found to be normal. Multiple myeloma are bone tumors similar to sar- coma, occurring usually in people beyond forty years of age, and in men more than in women. The condition is one of a deep-seated bone lesion asso- ciated with pain, emaciation, cachexia, etc. ; it is con- fined to the marrow and lives at the expense of the cancellous, tissue; it is limited to the long bones, the sternum and* ribs, rarely invoMng the skull. Bence-Jones protein has /been demonstrated in five types of cases : '"■■■■'■ 1. Multiple myeloma composed of (a) myeloblasts, (b) myelocytes, (c) erythroblasts. . 15 2. Chronic leukaemias (a) lympliatic. (bi niTelo- cytic. 3. Metastasis to bone from cancer elsewhere. 4. Other bone diseases present. 5. Cases without any lesions present. Usually not over two years elapse from the dis- covery of Bence-Jones protein to the death of the individual. In tAvo cases there was reported a high blood pressure. Xature of Bcncc-Joncs protein. It gives rise to amino acid partition and is there- fore a real protein, but it differs from any other known protein and is never found to be a part of the metabolism of the individual. It contains a car- bohydrate radicle, but no phosphorous, and is there- fore not a nucleo-protein. It also contains from 1 to 2 per cent of sulphur. It has been injected subcu- taneously, intravenously and intrarectally, but is al- ways excreted as such. Theories regarding Bence-Jones protein. 1. A substance derived from the secretory activity or degeneration of tumor masses themselves. 2. The tumor itself secretes an enzyme which can influence protein metabolism in such a manner that this protein is formed. 3. Individuals with Bejice-Jones protein have anomalies in urine excretion and metabolism similar to those excreting alkapton and cystin substances. It is probable that these individuals on account of their perverted metabolism are more prone to tumors. Character of the urine. The amount varies from 1200 to 2000 cc. per day with a normal specific gravity. It has a pale smoky color but is clear; it is acid in reaction, rarelv neu- 16 tral or alkaline ; it has a tendency to foam on shak- ing, which foam is more abundant and more per- sistent than normal. There may be as mncb as 70 grams of the protein present in the 24 honi speci- men, and there is no other type of protein nor any casts present unless there is some nephritic condi- tion superimposed. ClwracteriMics of the urine in this disease. 1. Continuous excretion regardless of everything. 2. Intermittent type, in which days or months may elapse without its excretion. 3. The appearance of the Bence-Jones protein sometimes antedates any demonstrable lesion. 4. The largest amounts are excreted in multiple myeloma which may run as high as 50 to 75 gm. per day. In other cases, such as metastatic cancer or sarcoma, from 20 to 35 gm. are excreted. 5. There is also a decreased chloride content. 6. A spontaneous precipitate may occur in the urine after standing for some time or it may occur in the bladder and give rise to a urine resembling chyle. Tests. 1. Heat and acetic acid. The urine clouds at about 55 degrees. It is coagulated and precipitafed from 10 to 20 degrees lower than any blood protein (40