A PRACTICAL TEXT- BOOK OF INFECTION, IMMUNITY AND SPECIFIC THERAPY WITH SPECIAL REFERENCE TO IMMUNOLOGIC TECHNIC BY JOHN A. KOLMER, M.D., DR.P.H., M.Sc. Assistant Professor of Experimental Pathology, University of Pennsylvania; Professor of Pathology and Bacteriology, Philadelphia Polyclinic, and Path- ologist to the Department of Dermatologic Research; Pathologist to the Philadelphia Hospital for Contagious Diseases WITH AN INTRODUCTION BY ALLEN J. SMITH, M.D., Sc.D., LL.D. Professor of Pathology, University of Pennsylvania WITH 147 ORIGINAL ILLUSTRATIONS, 46 IN COLORS By ERWIN F. FABER Instructor of Medical Drawing, University of Pennsylvania SECOND EDITION, THOROUGHLY RFV'SED PHILADELPHIA AND LONDON W. B. SAIJNDERS COMPANY 1917 LIBRARY Copyright, 1915, by W. B. Saunders Company. Reprinted July, 1915. Revised, reprinted, and recopyrighted October, 1917 Copyright, 1917, by W. B. Saunders Company PRINTED IN AMERICA PRESS OF W. B. SAUNDERS COMPANY PHILADELPHIA TO RICHARD M. PEARCE, M. D., D. SC. PROFESSOR OF RESEARCH MEDICINE IN THE UNIVERSITY OF PENNSYLVANIA IN RECOGNITION OF HIS SERVICES TO THE SCIENCE OF MEDICINE BY THE AUTHOR INTRODUCTION THE last quarter of a century has witnessed an almost marvelous development of knowledge in the domain of medicine and the allied sciences, only a part, of course, of the extensive progress made in the field of general science. A striking portion of this advance has tended to broaden our knowledge of the principles and of the essential details of the processes of infection and immunity, until these branches have today come to form almost a special science in themselves — an imperium in imperio. Aside from the personal factor, the writer's immediate inter- est in the present volume, as originally projected, arose from the fact that he was desirous of having appear a series of exercises illustrative of the principles of immunology — a class-book intended to set forth in' permanent form the very excellent course of instruction that Dr. Kolmer has been giving during the past few years to selected groups of in- terested students and occasional post-graduate workers in the Medical School of the University of Pennsylvania. That it should have sur- passed the original simple plan and grown into a volume of the present proportions is scarcely to be wondered at/ if the temptation to elaborate the individual exercises by explanations and cognate considerations was in the slightest to be yielded to. This is due to the fact that in its growth the subject has acquired so much of undoubted importance in the form of isolated observed facts, and itself presents so many analogies and has led to so extensive a terminology, that the author who would attempt to link the observed facts into anything like logical sequence or to add in the least to the bare cook-book-like series of illustrative exercises any explanatory paragraphs, cannot avoid the fullness that Dr. Kolmer has found inevitable in presenting the subject. The branch of immunology, including primarily infection, and its ramifications into diagnosis and the actual treatment of disease, has brought to the parent subject of preventive medicine the greatest offer- ing of the decades of its growth. Itself contributing to world expansion, it has nowhere found a greater stimulus than in the field of exotic pathol- ogy; and this last, in turn, has enriched internal medicine, even in its most common aspects. The first step in immunology may properly be ascribed to Jenner, with his bovine vaccine for smallpox, a step followed VI INTRODUCTION only after a long lapse of years by Pasteur. On the heels of the latter there appeared at once, and has since then followed, an army of men whose names crowd the history of the subject, and which many of these are bound permanently to adorn. The old vague theories of infection have taken form, and to observed facts has been added productive theory. The great danger attending this luxurious development is that, temporarily at least, the simpler, and perhaps the more obvious, facts are likely to be neglected; and, also, that symbolization by theories elaborated to harmonize with discovered facts will be accepted too fully as explanatory when in reality it does not explain, and that, as a result, investigation will finally be hampered instead of aided. In the almost universal drift of experimental studies to internal stereochemical factors, are we not in danger of placing too little stress upon actual and possible physical factors? Is there no danger that, by failing to lay stress upon the obvious importance of the turbinate mechanism in the nose as a natural anatomic factor, our rhinologists may at least feel justified in sacrificing this mechanism too readily for what may be but trivial local reasons? Can we insist that every phenomenon described with facility in terms of the side-chain theory is really a manifestation of chemism, when perhaps, with added investigation along lines of physical absorp- tion and the physical properties of colloids, an equally satisfying con- ception may be had, and possibly new facts be developed? Are we not blundering in rushing madly after matters of specificity as determined by antigen, when perhaps in reality we are confronted by potential and kinetic modifications due to peculiarities of diet or environmental cir- cumstances? The verity of phagocytosis is open to proof by observa- tion, and its variations are likewise to be demonstrated. Is the explana- tion of opsonins so convincing that merely the word itself is enough to satisfy the investigator? Infection and immunity constitute a definite chapter in pathologic science. The processes lack the dignity of a separate science only in that they present variations, and the fact that these are glossed over by brilliant theories and conceptions cannot prevent the deliberate recogni- tion of serious incompleteness. Yet this criticism can be applied to the growth of every branch of scientific knowledge. It in no wise militates against the right and the need for setting forth the subject in the light that, for the time, is afforded it. The importance of the criticism lies only in its acknowledgment, lest the subject as at present understood be accepted as fixed. With this danger obviated, and with all theories accepted for the time only as working theories, and their adoption not INTRODUCTION Vll urged to curtail investigations based on other views, their prosecution can be heartily applauded. This is the view that the writer believes that Dr. Kolmer has had in mind in his presentation of the subject as here set down. It is cer- tainly true of the chapters that the present writer has had opportunity of examining. In such a sense, therefore, the work is urged on the ap- preciation of the student, whether a laboratory worker or a mere seeker of knowledge. I have often been asked to what extent I believe it profitable to pre- sent the subject to the undergraduate student. I do not hesitate to answer that so far as the roster of the medical curriculum will permit, the laboratory demonstrations and exercises should form a part of the required course; and that, with all due caution to emphasize the fact that our present theory is not known to be final, and is offered merely tentatively, the verbal picture of the subject should be outlined before these beginners. To form some conception is necessary; and it is better, provided the mind be kept receptive, to follow a certain theory, even if it is unproved, than to do nothing at all or to work in confusion. Our American medical curriculum for undergraduates is so crowded with absolute essentials that the present subject is habitually neglected, save for a rapid lecture outline; this is an injustice to the student and to American medicine. I have tried to minimize this by providing, through Dr. Kolmer's aid, a reasonable laboratory course in the essentials of the branch to volunteer classes at first, at hours that did not interfere with the regular curriculum — at present during periods open to elec- tion. Nevertheless, the subject, influencing as it does every branch of medical practice, must take its place with other commendable additions to the required schedule. That this can be done only by lengthening the course of study, either in the annual session or by adding a year to our present four-year course, is obvious, and to that end we are rapidly approaching. ALLEN J. SMITH. PREFACE TO THE SECOND EDITION THE purpose and general plan of this volume remains unchanged in this edition. I have endeavored to make it a practical treatise for medical students, practitioners, and laboratory workers by setting forth theories and opinions simply and plainly, and devoting particular attention to technic and the practical phases and application of the subjects considered; I hope that this edition will receive the same gen- erous recognition as its predecessor. Additions and alterations have been made throughout; special attention has been given the subject of focal infection; the Schick toxin test for immunity in diphtheria and active immunization in diphtheria with toxin-antitoxin mixtures; complement-fixation in tuberculosis and other bacterial infections and a quantitative Wassermann reaction based upon my studies with the co-operation and assistance of Dr. Claude P. Brown, Dr. Toitsu Matsunami, and Dr. Berta Meine, aiming to standardize this important test. The chapters on anaphylaxis have been revised and particular attention given the subject of anaphylactic skin reactions. Lange's colloidal gold reaction has been included. The chapter on the treatment of various infections with bacterial vac- cines has been enlarged and the non-specific activity of bacterial vac- cines discussed. The section on the treatment of certain of the acute infectious diseases, and particularly acute anterior poliomyelitis, with the serum of convalescents and normal persons has been amplified; blood transfusion has been included. Special attention has been de- voted to the chapter on Chemotherapy, and the results of the studies of Dr. Jay F. Schamberg, Dr. George Raiziss, and the author bearing upon the toxicity of salvarsan and its congeners and the reactions following their administration have been included and discussed. The subject of Bacterial Chemotherapy, which promises much in the future, has been amplified from the theoretical and technical viewpoints. The section on Experimental Infection and Immunity remains as part of the volume instead of forming a separate book; experience has shown that this is a good plan, and enables the student doing more or less independent work to consult the text for directions and discussions. J. A. K. MCMANES LABORATORY OF EXPERIMENTAL PATHOLOGY, UNIVERSITY OF PENNSYLVANIA, October, 1917. ix PREFACE FOR the past twenty years the science of immunity has been one of the most progressive and most active branches in the department of medicine. An enormous literature has accumulated; many new terms have been coined, and numerous theories have been adduced; indeed, the subject has acquired an aspect of complexity that is confusing to those not specially interested or engaged in this work. The purpose of this book is a threefold one, namely : 1 . To give to practitioners and students of medicine a connected and con- cise account of our present knowledge regarding the manner in which the body may become infected, and the. method, in turn, by which the organism serves to protect itself against infection, or strives to overcome the infection if it should occur, and also to present a practical application of this knowledge to the diagnosis, prevention, and treatment of disease. 2. To give to physicians engaged in laboratory work and special workers in this field a book to serve as a guide to the various immunologic methods. 3. To outline a laboratory course in experimental infection and im- munity for students of medicine and those especially interested in these branches. 1. The subject of infection is intimately connected with that of immunity, and this is especially emphasized in those diseases for which a specific therapy exists, for a knowledge of the nature of the infection is of paramount importance in controlling the dosage and indicating the method of administration of a specific therapeutic agent. By describing principles and technic with considerable detail, a special effort has been made to render Part IV of this book of particular value to practitioners of medicine. The day is past when the physician and surgeon can relegate the things of immunity entirely to the laboratory. Diagnostic methods and re- actions and the field of specific therapy, — vaccine, serum, and chemo-, — are subjects of such practical importance that it is obvious that the physician and the student of medicine can no longer be merely mildly interested onlookers. The physician who injects salvarsan, a serum, or a vaccine, or who uses a diagnostic reaction, must be prepared to ex- plain to his patient the nature of the therapy he employs and the sig- Xll PREFACE nificance of the reaction. This he can do only by equipping himself with the knowledge of the fundamental factors of immunity, or he will be forced into the position of a passive transmitter of ideas entirely be- yond his own knowledge. 2. An effort has been made to include data of both practical and theoretic importance, and in some instances tests are described that are more of theoretic than of practical import, especially in research work. » It is obviously impossible, in a single volume, to include the very large number of tests and modifications that have been advocated from time to time, and, as a matter of course, most attention has been given those methods that have been shown to be of practical value or that give promise of becoming so. So far as possible original methods are given, these being, in the larger proportion, more or less important modi- fications devised as the result of my own experience in hospital and teach- ing laboratories. The technic of the various tests and reactions is described in great detail, thus tending the better to secure accuracy, simplicity, and definite- ness, and to serve as an opening wedge to those about to enter this special field. 3. The value of the experimental method in the teaching of certain branches of medicine is now well recognized. In no department, how- ever, is this method of greater value than in the study of infection and immunity. A working knowledge of these subjects is so valuable in the practice of medicine and surgery that the student should be well versed in at least their primary principles and practical applications in the prophylaxis, diagnosis, and treatment of diseas The laboratory course given in Part V is based upon the courses given by me in the Laboratory of Experimental Pathology at the Uni- versity of Pennsylvania, and in the laboratories of the Philadelphia Polyclinic and College for Graduates in Medicine. In including them in this volume I am carrying out my original plan, for in many of the experiments the exact technic of a given test is described, making a sepa- rate book devoted to this part of the subject unnecessary. Future ex- perience may, however, show the necessity of having this portion of the book form a separate laboratory manual. I shall appreciate the opin- ions of educators who may have occasion to consult the course herein outlined. Since the larger portion of our knowledge of infection and immunity has been gained from studies upon the lower animals, it is not strange that these were early and directly benefited by a practical application PREFACE Xlii of this knowledge to the prophylaxis, diagnosis, and treatment of many of the diseases to which these animals are subject. I have, therefore, included in this volume an account of those immunologic diagnostic reactions and applications of specific therapy that have a direct bearing upon veterinary medicine. No attempt has been made to cover all literature references on the subject. An effort has been made to state well-established facts con- cisely, and, in the case of the more recent subjects, to give the principal references to the literature. I have drawn largely from German, French, and English sources, and have endeavored, wherever possible, to give proper and due credit to each author. In order to keep the work up to the times, I would ask the authors of reprints on immunologic subjects to send me copies. The illustrations, all of which have been made by Mr. Erwin F. Faber, will, it is hoped, serve the purpose for which they are intended, namely, to elucidate the text and to teach, rather than merely to em- bellish. It is with deep and sincere appreciation that I acknowledge the en- couragement and aid given me by Professor Allen J. Smith, who has written the introduction and reviewed several chapters. My thanks are also due to Professor Richard M. Pearce for reviewing the chapters on Infection; to my assistant, Dr. Anna M. Raiziss, for a number of translations from foreign literature, and to the publishers, whose kind and unvarying courtesy has greatly simplified the work. J. A. K. MCMANES LABORATORY OF EXPERIMENTAL PATHOLOGY, UNIVERSITY OF PENNSYLVANIA. CONTENTS PAGE INTRODUCTION v PART I GENERAL IMMUNOLOGIC TECHNIC CHAPTER I. — GENERAL TECHNIC 17 Care of centrifuge, 17 — Making a simple capillary pipet, 18 — Making looped pipets, 20 — Graduated pipets, 21 — Making Wright blood-capsules, -23 — Making vaccine ampules, 24 — Preparation of test-tubes for immunologic work, 25 — Selection of a satisfactory syringe, 26 — Solutions, 27. CHAPTER II. — METHODS OF OBTAINING HUMAN AND ANIMAL BLOOD 28 Obtaining corpuscles, 28 — Washing erythrpcytes, 28 — Obtaining serum, 30 — Obtaining corpuscles and serum, 30 — Obtaining blood plasma, 30— Obtaining small amounts of human blood, 32 — Obtaining larger amounts of human blood (phlebotomy, wet-cupping, placental blood), 33 — Obtaining cerebrospinal fluid (technic of spinal puncture), 37 — Obtaining small amounts of animal blood (rabbit, guinea-pig, sheep), 41 — Obtaining large amounts of animal blood (rab- bit, guinea-pig, rat, sheep, hog, monkey, dog, and horse), 42. CHAPTER III. — TECHNIC OF ANIMAL INOCULATION 53 General rules, 53 — Method of subcutaneous inoculation (fluid and solid inocula), 54 — Method of intramuscular inoculation, 56 — Methods of intravenous inoculation (rabbit, guinea-pig, mice and rats, horse, sheep, goat, dog), 56 — Method of intracardial inoculation, 62 — Methods of intraperitoneal inoculation (rabbit, guinea-pig), 64. CHAPTER IV. — METHODS FOR EFFECTING ACTIVE IMMUNIZATION OF ANIMALS . . 65 Antigens and active immunization, 65 — General technic, 66 — Production of antitoxins, 68 — Production of agglutinins (intravenous and intraperitoneal inocu- lation), 68 — Production of immune opsonins, 69 — Production of bacteriolysins, 69 — Production of precipitins, 70 — Production of hemolysins (intravenous and intraperitoneal inoculations), 71 — Production of cytotoxins, 73. CHAPTER V. — PRESERVATION OF SERUMS — METHODS 75 Methods for the preservation of normal serums, 75 — Methods for the preser- vation of immune serum in fluid form with antiseptics, 76 — In fluid form by bac- teria-free filtrations, 76 — In fluid form by freezing, 79 — Preservation in powder form, 79 — Preservation in dried paper form, 80. PART II PRINCIPLES OF INFECTION CHAPTER VI. —INFECTION 81 Definition, 81 — Relation of infection to immunity, 83 — Source of infection, 83 — Contagious and infectious diseases, 84 — Exogenous and endogenous in- fection, 85— -Avenues of infection, 86 — Normal defenses against bacterial in- vasion, 90 — Mechanism of bacterial invasion, 91 — Local infection, 94 — Mechan- ism of infection, 94 — The avenue of infection and tissue susceptibility, 98 — The numeric relationship of bacteria to infection, 99 — General susceptibility in relation to infection, 100 — The defensive mechanism of the microorganism in relation to infection, 103 — Mixed infection, 106 — Summary, 107. 4 CONTENTS PAGE CHAPTER VII. — INFECTION (continued). PRODUCTION OF DISEASE 108 Toxins, 109 — Extracellular bacterial toxins, 110 — General properties of soluble toxins, 110— Precipitation of the extracellular toxins, 111 — Struc- ture of soluble toxins, 111 — Nature of soluble toxins, 112 — Selective action of soluble toxins, 112. Special Properties of the Principal Soluble Toxins, 113 — Diphtheria toxin, 113 — The guinea-pig test for virulence of diphtheria bacilli, 114 — Tetanus toxin, 117 — Botulism toxin, 117 — Dysentery toxin, 118 — Staphylotoxin, 118— Streptotoxin, 119. Toxins of the Higher Plants and Animals, 119 — Phytotoxins, 119 — Pollen toxin, 120— Zootoxins, 121 — Snake venom, 121. Endotoxins, 122 — Methods of studying endotoxins, 122 — Nature of endotoxins, 123 — Aggressins, 124 — Bail's classification of bacteria, 125 — Nature of aggressins, 126 — Anti-aggressins, 127 — .Bacterial Proteins, 127 — Bacterial split protein, 128 — Nature of bacterial proteins, 128 — Action of bac- terial proteins, 129 — Theory of Vaughan, 130. Ptomains, 131. Mechanical Action of Bacteria, 133 — Infection with Animal Parasites, 134 — The Course of Infection, 136 — Stages of infection, 136 — Grades of Infection, 138 — Systemic reaction to infection, 139. PART III PRINCIPLES OF IMMUNITY AND SPECIAL IMMUNOLOGIC TECHNIC CHAPTER VIII. — IMMUNITY. — THEORIES OF IMMUNITY 140' Definition, 142 — Historic, 142 — Exhaustion theory of immunity, 143 — Retention theory of immunity, 146 — Theory of phagocytosis, 146 — Side-chain theory of immunity, 148 — Compatibility of the phagocytic and side-chain theories, 157 — Antigens, 161 — Antibodies, 163. CHAPTER IX. — VARIOUS TYPES OF IMMUNITY 167 Natural Immunity, 167 — Explanation of natural immunity, 168 — Non- specific immunity, 168 — Local immunity, 169 — Phagocytosis and natural im- munity, 170— Natural antitoxic immunity, 171 — Natural bacteriolytic immun- ity, 171 — Natural anti-aggressin immunity, 171 — Athreptic immunity, 172. Acquired Immunity, 172 — Active acquired immunity, 172 — Passive acquired im- munity, 174. CHAPTER X. — PHAGOCYTOSIS 177 Historic, 177 — The original theory of phagocytosis, 178— Kinds of phagocy- tosis, 179 — The relation of the cell types to infection, 180 — Chemotaxis, 181 — Positive chemotaxis, 181 — Negative chemotaxis, 184 — Results of phagocytosis, 184 — The relation of body fluids to phagocytosis, 186 — Revised theory and role of phagocytosis in immunity, 188 — Mechanism of phagocytosis, 189. CHAPTER XI. — OPSONINS 190 Historic, 190 — Definition, 191— Properties and nature of opsonins, 191— Susceptibility to opsonification, 192— Effect of opsonins on bacteria, 192— Role of opsonins in immunity, 193. CHAPTER XII. — OPSONIC INDEX 194 Principles involved, 194 — Definition, 194 — Purpose of the opsonic index, 195— Limitations of the method, 195 — Precautions in technic, 196— Technic of the opsonic index (Wright), 196 — Technic of quantitative estimation of bacterio- tropins in immune serum (Neufeld), 203 — Practical value of the opsonic index, 206 — Value of the index in diagnosis, 207 — In prognosis, 208 — As a guide to bacterial vaccine therapy, 208. CHAPTER XIII. — BACTERIAL VACCINES 210 Definition, 210 — Technic of preparing bacterial vaccines, 210— Counting a bacterial vaccine (method of Wright), 214 — Counting with the hemocytometer chamber, 215— Method of Kolle, 216— Method of Hopkins, 216— Preparation of "sensitized" bacterial vaccine, 221 — The administration of a bacterial vaccine, 221 — Making the inoculation, 221 — Effects of inoculation, 222— Frequency and dosage of inoculation, 222 — Ordinary adult doses of the common vaccines, 222. CONTENTS 5 PAGE CHAPTER XIV. — ANTITOXINS 224 Definition, 224 — Historic, 224 — Formation of antitoxins, 225 — Structure of antitoxins, 227 — Properties of antitoxins, 227 — Natural antitoxins, 228 — Schick test for natural diphtheria antitoxin, 228 — Specificity of antitoxins, 232 — Nature of the toxin-antitoxin reaction, 232 — Production of Diphtheria Anti- toxin, 235 — Production of diphtheria toxin, 235 — Testing the toxin, 236 — Im- munizing the animals, 236 — Collecting the serum, 239 — Method of concentrat- ing, 239— Standardizing the serum, 240. Production of Tetanus Antitoxin, 243 —Tetanus toxin, 243 — Immunizing the animals, 244 — Collecting the serum, 244 — Standardizing the serum, 244. Botulimis Antitoxin, 245. Antidysentery Serum, 245 — The culture, 246 — Immunizing the animals, 247 — Rapid produc- tion, 247 — Collecting and testing the serum, 248. Antistaphylococcus Serum, 249 — Preparation, 249 — Technic of the anti-lysin test, 249. Production of Antivenin, 251. Production of Pollen Antitoxin, 252. The Measure of Anti- toxins, 252 — A unit, 252 — Unit of diphtheria antitoxin, 253 — Unit of tetanus antitoxin, 253. CHAPTER XV. — FERMENTS AND ANTIFERMENTS 254 Bacterial ferments, 254 — Similarity between toxins and ferments, 254 — Antif erments, 256 — Antibodies and antiferments, 257 — Antiferments in disease, 257 — Ferments in pregnancy and disease, 258. Mechanism of the Abderhalden reaction, 260. Ferment Reactions, 264— Antitrypsin test, 264— Abderhalden 's serodiagnosis of pregnancy, 265 — Practical value of Abderhalden's test, 276 — Sero-enzymes in cancer, 277 — In mental diseases, 278 — In syphilis, 278 — In tuberculosis and acute infection, 278. CHAPTER XVI. — AGGLUTININS 279 Definition, 279 — Historic, 279— Normal and immune agglutinins, 281 — Formation of agglutinins, 281 — Origin of agglutinins, 282 — Properties and nature of agglutinins, 283 — Acid agglutination, 283 — Mechanism of agglutina- tion, 284 — Specificity of agglutinins, 285 — Absorption methods for differentiat- ing between a mixed and single infection, 285 — Hemagglutinins, 286 — Non-ag- glutinable species of bacteria, 288 — Variation in agglutinating strength of a serum, 288 — Conglutination, 289 — Role of agglutinins in immunity, 289. Practical Applications, 290 — In the diagnosis of typhoid fever, 290 — Paraty- phoid fever, 291 — Dysentery, 292 — Cholera, 292— Cerebrospinal meningitis, 292— Plague, 292— Malta fever, 292— Pneumonia, 292— Syphilis, 292— Per- tussis, 293 — Glanders, 293 — In the differentiation of bacteria, 293 — In the diag- nosis of single and mixed infection, 294. The Agglutination Reaction, 294— Microscopic method with serum, 290 — Microscopic method with dried blood, 300 — Macroscopic method, 301 — Differentiation of pneumococci, 308 — The saturation test of Castellani, 309 — Technic of acid agglutination, 310. Tests before Blood Transfusion for Isohemagglutinins and Isohemolysins, 311 — Microscopic method of Brem, 312. CHAPTER XVII. — PRECIPITINS 314 Definition, 314 — Historic, 314 — Nomenclature, 316 — Structure and propor- tion of precipitins, 316 — Formation of precipitins, 317 — Mechanism of precipita- tion, 318 — Specificity of precipitins, 318 — Role of precipitins in immunity, 319. Practical Applications, 320 — Bacterial precipitins, 320 — Fornet ring test, 321 — Porges-Meier reaction, 321 — Herman-Perutz reaction, 322— Noguchi globulin re- action, 322 — McDonagh ''gel" test, 323 — Differentiation of proteins, 324. Technic of the Precipitin Reactions, 326 — Differentiation of human and animal bloods, 326 — Detection of meat adulteration, 333 — Bacterial precipitins, 336. CHAPTER XVIII. — CYTOLYSINS. AMBOCEPTORS AND COMPLEMENTS 338 Definition, 339 — Kinds of cytolysins, 340 — Nomenclature, 340. Ambocep- tors, 341 — Historic, 341 — Structure of amboceptors, 342 — General properties of amboceptors, 343 — Mechanism of the action of amboceptors, 343 — Formation of amboceptors, 344 — Quantitative estimation of amboceptors, 347 — Titration of hemolytic amboceptor, 347 — Titration of bacteriolytic amboceptor, 347. Complements, 348 — Historic, 348 — Definition, 348 — Structure and general properties of complement, 348 — Anticomplements, 349— Origin of complements, 350 — Multiplicity of complements, 350 — Endocomplements, 352 — Complement- splitting, 353 — Complement fixation, 354 — Complement deviation, 355 — Quan- titative titration of complement, 356. D CONTENTS CHAPTER XIX. — BACTERIOLYSINS 358 Historic, 358 — Definition, 359 — Origin of bacteriolysins, 360 — Leukins and leukocytic extracts, 360 — Method of preparing leukocytic extracts, 361 — Mech- anism of bacteriolysis, 362 — General properties of bacteriolysins, 363 — Normal bacteriolysins, 363 — Specificity of bacteriolysins, 363. Practical Applications, 363— Technic of the Pfeiffer test, 364 — Bacteriolytic test in vivo f9r the identi- fication of bacteria, 365 — Bacteriolytic test in vivo in the diagnosis of disease, 370 — Bacteriolytic test in vitro (method of Stem and Korte), 371 — Bacterio- lytic test in vitro (method of Wright), 377. CHAPTER XX. — HEMOLYSINS 383 Historic, 383 — Definition, 384 — Nomenclature, 385 — Nature of hemolysins, 385 — Analogy between bacteriolysis and hemolysis, 389 — Specificity of hemoly- sins, 390 — Normal hemolysins, 390 — Production of immune hemolysins, 393 — General properties of hemolysins, 393 — Source of hemolysins, 394. Practical Applications, 395 — Quantitative reactions between hemolytic amboceptor and complement, 396 — Method of titration of hemolysin, 397 — Method for removing hemolysin from a serum, 400 — Method of determining natural hemolysins in serum, 400-^The serum diagnosis of paroxysmal hemoglobinuria, 401 — Method of determining the resistance of red blood-corpuscles, 402. CHAPTER XXI. — VENOM HEMOLYSIS : 405 Historic — nature of venom hemolysis, 405— Venom hemolysis in syphilis, 407 — Practical value of the venom test in syphilis, 410 — The psycho-reaction of Much in syphilis, 410 — Venom hemolysis in tuberculosis, 412 — Venom hemolysis in cancer, 412. CHAPTER XXII. — PRINCIPLES OF THE PHENOMENON OF COMPLEMENT FIXATION 413 Historic, 413 — The original complement-fixation method of Bordet, 416 — Mechanism of complement fixation, 417 — Non-specific complement fixation, 418 — Quantitative factors in complement-fixation tests, 421 — Practical applica- tions, 423. CHAPTER XXIII. — THE TECHNIC OF COMPLEMENT-FIXATION REACTIONS 425 The Wassermann Syphilis Reaction, 425 — Historic, 425 — Principles and theories of the syphilis reaction, 428 — General technic, 431 — Preparation of the fluid to be tested, 432 — Preparation and titration of complement, 435— Preserva- tion of complement, 437 — Preparation and titration of hemolytic amboceptor, 439 — Preparation of blood-corpuscles, 440 — Preparation and standardization of antigens, 441— Technic of the First Method, 459— Technic of the Second Method, 465-^Technic of the Third Method, 467 — Technic of the Fourth Method, 470. Modifications of the Wassermann Reaction, 475 — Technic of the Noguchi modi- fication, 476 — Technic of the Hecht-Gradwohl modification, 482 — Other modi- fications, 484. The Wassermann Reactions in the Various Stages of Syphilis, 485 — The specificity of the Wassermann reaction, 490 — The effect of treatment upon the Wassermann reaction, 492 — The Practical Value of the Wassermann reaction, 495. CHAPTER XXIV. — COMPLEMENT-FIXATION REACTIONS (continued) 498 Specific complement fixation in bacterial diseases, 498 — Preparation of bacterial antigens, 498 — Standardizing bacterial antigens, 500 — Principles of complement fixation with bacterial antigens, 501. Complement Fixation in the Differentiation of Microparasites, 502. Complement Fixation in Gonococcus Infections, 503. Complement Fixation in Glanders, 511. Complement Fixa- tion in Contagious Abortion, 513. Complement Fixation in Dourine, 515. Complement Fixation in Typhoid Fever, 516. Complement Fixation in Tuber- culosis, 517. Complement Fixation in Pertussis, 521. Complement Fixation in the Standardization of Immune Serums, 524. Complement Fixation in Echinococcus Disease, 524. Complement Fixation in the Differentiation of Proteins (blood-stains, meats, bacteria), 527. Complement Fixation in Cancer, 532. CONTENTS 7 PAQK CHAPTER XXV.— CYTOTOXINS 534 Nomenclature, 534 — Nature and general properties of cytotoxins, 534 — Preparation of cytotoxins, 535 — Methods of studying cytotoxins, 535 — Specific- ity of cytotoxins, 536 — Autocytotoxins, 537 — Isocytotoxins, 538 — Anticytotoxic serums, 538 — Kinds of cytotoxins, 538 — Spermatotoxin, 538 — Epitheliotoxin, 538 — Leukotoxin, 538 — Nephrotoxin, 538 — Hepatotoxin, 539 — Gastrotoxin, 539 — Syncytotoxin, 539 — Neurotoxin, 540 — Thyrotoxin, 540 — Role of cyto- toxins in immunity, 540 — Practical applications, 540 — Cytotoxic cancer reac- tion, 541. CHAPTER XXVI. — THE RELATION OF COLLOIDS AND LIPOIDS TO IMMUNITY . . . 543 Kinds of colloids, 543 — Nature and properties of colloids, 543 — Analogy be- tween the reactions of immunity and those of colloidal chemistry, 548 — Anti- toxins, 548 — Agglutinins and precipitins, 55fr— Hemolysins, 551 — Complement fixation as a colloid reaction, 552 — The relation of lipoids to immunity, 553 — Lange's colloidal gold reaction, 555 — The epiphanin reaction, 559 — The miostag- min reaction, 563. CHAPTER XXVII. — ANAPHYLAXIS 567 Historic, 568 — Definition, 571 — Phenomena of anaphylaxis, 572 — Mechan- ism of anaphylaxis, 577 — Anaphylactogens or allergens, 579 — The humeral or anaphylotoxin theories of anaphylaxis, 584 — Anaphylactin (allergin), 588 — Theories of anaphylaxis, 591 — Cellular theory of anaphylaxis, 594r— Passive anaphylaxis, 596 — Anti-anaphylaxis or desensitization, 599 — Specificity of anaphylaxis, 601. PART IV APPLIED IMMUNITY IN THE PROPHYLAXIS, DIAGNOSIS, AND TREATMENT OF DISEASE— SPECIFIC THERAPY CHAPTER XXVIII. — ANAPHYLAXIS IN ITS RELATION TO INFECTION AND IM- MUNITY. ANAPHYLACTIC OR ALLERGIC REACTIONS 602 Relation of anaphylaxis to infectious diseases, 603 — Relation of anaphylaxis to non-infectious diseases, 608 — Serum disease, 608 — Idiosyncrasies, 614 — Relation of anaphylaxis to immunity, 618 — Anaphylactic or allergic skin reac- tions, 620 — Subcutaneous tuberculin reaction, 633 — Intracutaneous tuberculin reaction, 636 — Cutaneous tuberculin reaction, 637 — Conjunctival tuberculin re- action, 639 — Percutaneous tuberculin reaction, 640— Tuberculin reactions among the lower animals, 641 — The luetin reaction, 642 — The mallein reaction, 647 — Abortin reaction, 648— Allergic reactions in typhoid fever, 649 — Allergic reactions in other diseases, 652. CHAPTER XXIX.— ACTIVE IMMUNIZATION. VACCINES IN THE PROPHYLAXIS AND TREATMENT OF DISEASE. VACCINE THERAPY 654 Historic, 654 — Nomenclature, 656 — Method of preparing vaccines, 657 — Mechanism of active immunization, 659 — Non-specific activity, 663 — Living versus dead vaccines, 664 — Sensitized vaccines, 665 — Autogenous versus stock bacterial vaccines, 665 — The negative phase, 666 — Contraindications to active immunization, 667. Prophylactic Irtimunization or Vaccination, 668 — In small- ?ox, 668 — In rabies, 681 — In typhoid fever, 688 — In paratyphoid fever, 693 — n typhus fever, 694 — In pneumonia, 694 — In plague, 694 — In cholera, 697 — In dysentery, 699 — In cerebrospinal meningitis, 699 — In scarlet fever, 699 — In pertussis, 700 — In anthrax, 700 — In black-leg, 702. Therapeutic Immunization, Bacterial Vaccine Therapy, 702— Principles, 702— Infected wounds, 703 — Focal infections, 704 — Diseases of the skin, 704 — Diseases of the genito-urinary sys- tem, 706 — Diseases of the respiratory system, 708 — Acute general infections, 713— Tuberculin therapy, 713. QE i 8 CONTENTS PAGE CHAPTER XXX. — PASSIVE IMMUNIZATION. SERUMS IN THE PROPHYLAXIS AND TREATMENT OF DISEASE. SERUM THERAPY 73 Definition, 733— Purposes of passive immunization, 734 — Kinds of passive immunity, 735 — Indications for passive immunization, 736 — Contraindications to passive immunization, 738 — Technic of subcutaneous inoculation, 740 — Technic of intramuscular inoculation, 742 — Technic of intravenous inoculation, 742 — Technic of subdural inoculation, 746. Serum Treatment and Prophylaxis of Diphtheria, 754. Serum Treatment and Prophylaxis of Tetanus, 772. Se- rum Treatment and Prophylaxis of Dysentery, 782. Serum Treatment and Prophylaxis of Hog Cholera, 784. Serum Treatment of Snake Bites, 786. Serum Treatment and Prophylaxis of Hay-fever, 786. Serum Treatment and Prophylaxis of Meningococcus Meningitis, 788. Serum Treatment of Influenzal Meningitis, 802. Serum Treatment of Pneumococcus Meningitis, 804. Serum Treatment of Other Localized Pneumococcus Infections, 807. Serum Treat- ment of Lobar Pneumonia, 807. Serum Treatment of Streptococcus Infections, 813. Serum Treatment of Gonococcus Infections, 818. Serum Treatment of Staphylococcus Infections, 819. Serum Treatment of Anthrax, 820. Serum Treatment of Typhoid Fever, 821. Serum Treatment of Plague, 822. Serum Treatment of Cholera, 823. Serum Treatment of Tuberculosis, 824. Treatment of Infectious Diseases with Specific Serum of Convalescents and with Normal Serum, 825. Serum Treatment of Scarlet Fever, 825. Serum Treatment of Acute Anterior Poliomyelitis, 826. Serum Treatment of Pneumonia, 828. CHAPTER XXXI. — SERUM THERAPY (continued) 829 Normal Serum Therapy, 829 — Serum treatment of hemorrhage, 829— ^Serum treatment of the toxicoses of pregnancy, 833 — Serum treatment of skin diseases, 834. Autoserum Therapy, 835— Autoserum treatment of skin diseases, 835 — Autoserum treatment of acute infectious diseases, 835 — Autoserum treatment of syphilis (salvarsanized serum), 836 — Autoserum treatment of tuberculosis of serous membranes, 841 — Autoserum treatment of non-tuberculous effusions, 843. CHAPTER XXXII. — CHEMOTHERAPY 844 Principles of Chemotherapy, 845 — Organotropism and parasitotropism, 845 — Chemoreceptors, 847 — Drug "fastness," 849— Therapia magna sterilisans, 851. Salvarsan and Neosalvarsan in the Treatment of Syphilis, 852 — Historic, 852 — Properties of salvarsan, 854 — Toxicity of .salvarsan, 855 — Properties of neo- salvarsan, 856— Methods for preparing salvarsan for administration-, 857 — Methods for preparing neosalvarsan for administration, 857 — Administration of salvarsan and neosalvarsan by intravenous injec^n, 858 — Reactive manifesta- tions, 867 — By intramuscular injection, 875 — By intraspinous injection, 876 — Contraindications and precautions in salvarsan therapy, 878 — Value of salvarsan and neosalvarsan in the treatment of syphilis, 879. Salvarsan in the Treatment of Non-syphilitic Diseases, 880. Chemotherapy in Bacterial Diseases, 881. Chemotherapy in Malignant Diseases, 887. PART V EXPERIMENTAL INFECTION AND IMMUNITY Introductory, 890 — Methods, 890 — The Student, 890 — Records, 891— Animal Experiments and Autopsies, 891. EXERCISE 1. — Active Immunization of Animals, 891. Experiment 1, Production of Agglutinins, .Bacteriolysins and Opsonins, 891 — Experiment 2, Production of Precipitins, 892— Experiment 3, Production of Hemolysins, 892 — Experiment 4, Production of Cytotoxin, 892. EXERCISE 2. — Infection, 892. Experiment 5, Experimental Pneumonia, 892. EXERCISE 3. — Toxins, 893. Experiment 6, Diphtheria Toxin, 893 — Experiment 7, Method of Testing the Virulence and Toxicity of Diphtheria Bacilli, 894— Experiment 8, Tetanus Toxin (in vivo}, 895 — Experiment 9, Tetanus Toxin (in vitro), 895. CONTENTS 9 EXERCISE 4. — Toxins (continued), 896. Experiment 10, Botulism Toxin, 896— Experiment 11, Dysentery Toxin, 896— Experiment 12, Staphylotoxin (in vivo), 897 — Experiment 13, Staphylotoxin (in vitro), 897. EXERCISE 5. — Toxins (continued) — Plant and Animal Toxins, 898. Experiment 14, Streptotoxin, 898 — Experiment 15, Phytotoxins (in vivo and in vitro), 898 — Experiment 16, Zootoxin, Cobra venom (in vivo and in vitro), 899. EXERCISE 6. — Endotoxins and Aggressins, 899. Experiment 17, Endotoxins, 899 — Experiment 18, Natural Aggressins, 900. EXERCISE 7. — Bacterial Protein. Ptomains. Mechanical Action of Bacteria, 900. Experiment 19, Bacterial Protein, 900 — Experiment 20, Ptomains, 901 — Ex- periment 21, Mechanical Action of Bacteria, 902. EXERCISE 8. — Kinds of Immunity. Natural Immunity, 902. Experiment 22, Phagocytosis in Natural Immunity, 902 — Experiment 23, Nat- ural Antibacterial Immunity, 902 — Experiment 24, Relative Factors in Natural Immunity, 903 — Experiment 25, Influence of Temperature Upon Natural Immunity, 903. EXERCISE 9. — Acquired Immunity, 903. Experiment 26, Acquired Active (Antibacterial) Immunity, 903 — Experiment 27, Acquired Passive (Antitoxic) Immunity, 904 — Experiment 28, Acquired Passive (Antitoxic) Immunity, 904 — Experiment 29, Acquired Passive (Anti- bacterial) Immunity, 904. EXERCISE 10. — Phagocytosis, 905. Experiment 30, Phagocytosis (macrophages), 905— Experiment 31, Phagocy- tosis (microphages), 905 — Experiment 32, Phagocytosis, 906. EXERCISE 11. — Phagocytosis. Chemotaxis, 906. Experiment 33, Positive Chemotaxis, 906 — Experiment 34, Negative Chemo- taxis, 906. EXERCISE 12. — Opsonins, 907. Experiment 35, Normal Opsonins, 907 — Experiment 36, Immune Opsonin (Bac- teriotropin), 908 — Experiment 37, Hemopsonin, 908. EXERCISE 13. — Opsonins (continued), 908 Experiment 38, Mechanism of Action of Opsonins, 908 — Experiment 39, Specific- ity of Opsonins, 909. EXERCISE 14. — Opsonic Index, 910. Experiment 40, Determining the Opsonic Index, 910 — Experiment 41, Quan- titative Estimation of Bacteriotropins, 910. EXERCISE 15. — Bacterial Vaccines, 910. Experiment 42, Preparation of Typhoid Vaccine, 910 — Experiment 43, Prepara- tion of Staphylococcus Vaccine, 911. EXERCISE 16. — Antitoxins, 911. Experiment 44, Standardizing Diphtheria Antitoxin, 911 — Experiment 45, Standardizing Tetanus Antitoxin, 912. EXERCISE 17. — Antitoxins (continued), 912. Experiment 46, Specificity of Antitoxins, 912 — Experiment 47, Nature of the Toxin-Antitoxin Reaction. Action of Anti-tetanolysin, 912 — Experiment 48, Antistaphylolysin, 913. EXERCISE 18. — Ferments and Antiferments, 914. Experiment 49, Tryptic Ferment of Leukocytes, 914 — Experiment 50, Testing the Antitryptic Power of Bl