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The distribution and storage of blue antigenic azoproteins in the tissue of mice

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Abstract

Intensely blue dye-azoproteins have been prepared by diazotization and coupling of the highly indiffusible blue dye T-1824, Evans blue, with various serum proteins and egg albumin. The products, whether purified by precipitation with alcohol or by chromatography, have a constant dye-to-protein ratio and tests have shown them to be essentially free from unlinked dye. An extremely diffusible dye, echt-säure-blau, has also been coupled to bovine gamma-globulin. These materials are adapted to physiological experimentation. They seem to behave in the bodies of mice like other proteins; they fail to appear in either the bile or urine of normal animals, and they are strongly antigenic. When these soluble antigenic azoproteins are injected into the blood stream of mice for the first time they enter reticulo-endothelial cells in almost every organ of the body; the final distribution is like that of intravenously injected, finely divided particulate matter. The azoproteins appear in the cells which classical immunological studies have shown to be active in removing particulate antigenic materials or bacteria from the blood or body fluids. The Kupffer cells of the liver and sinus and reticular cells in lymph nodes, especially the great mesenteric node, are particularly active in the removal of the blue antigens from the blood, but many other R-E cells are active to a lesser degree. The storage of the antigenic material is in the cytoplasm only; it has not been seen within nuclei, nor has it been seen within cells of the brain. Serological methods disclose that the blue material seen within Kupffer cells of the liver after as long a period as 2 days is still antigenic in its reactions. The blue azoproteins, therefore, serve excellently as tracer antigens, especially since they can be seen directly in fresh and fixed tissue preparations and in the body fluids.

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... A more commonly used label, fluorescein isothiocyanate (Klatzo, Miquel & Otenaser, 1962), was found in pilot studies to be unsuitable, since its emitted fluorescence was difficult to distinguish from the autofluorescence of the sections, even when the most highly selective filters available were used. Evans blue, another fluorescent dye popular in studies of this type (Olsson & Kristensson, 1973;Vise et al. 1975), was not used because it does not bind covalently to serum proteins and even when covalent binding is deliberately induced, by azo-coupling, the chromogenic moiety can still be detached and translocated in vivo (Kruse & McMaster, 1949). ...
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Vital staining was studied by quantitating the amounts of Evans blue and serum albumin in muscle and in abnormal brain. On the basis of differences in the ratio of dye to albumin in tissue and in serum, it was concluded that the coloration of tissue involves both the presence of serum albumin in tissue and the binding of dye not associated with serum albumin by tissue. The mode of binding of free dye by edematous cerebral tissue was studied by comparing the chemical structures of a series of acid dyes which did and did not function as vital stains. It was concluded that substantivity for edematous cerebral tissue requires the presence on the dye molecule of paired widely separated electron-donating hydrogen bond forming groups placed away from the solubilizing acid groups. It is suggested that the primary mode of binding for Evans blue in tissue is through the formation of hydrogen bonds between the azo groups of the dye and the hydroxy groups of tissue polysaccharides. These findings are conceptually related to the blood-brain barrier in that the presence of staining in the brain reflects the presence of increased amounts of albumin either at the time of observation or at some time in the past.
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In order to reduce the extrahepatic side effects of antiviral nucleoside analogues in the treatment of chronic viral hepatitis, these drugs are conjugated with galactosyl-terminating macromolecules. The conjugates selectively enter hepatocytes after interaction of the carrier galactose residues with a receptor present in large amounts and high affinity only on these cells. Within hepatocytes the conjugates are delivered to lysosomes where enzymes split the bond between the carrier and the drug, allowing the latter to become concentrated in the liver. The majority of experiments referred to in this paper were performed employing a conjugate of lactosaminated human albumin with adenine arabinoside monophosphate (ara-AMP), a phosphorylated nucleoside analogue active against hepatitis B virus. This conjugate administered for 28 days to patients with chronic hepatitis B exerted the same antiviral activity as that of the free drug without producing any clinical side effects, including the severe neurotoxicity caused by free ara-AMP. This result demonstrates the validity of the liver targeting approach which enhances the therapeutic possibilities of nucleoside analogues. Coupling to galactosyl terminating carriers may be a way of obtaining higher drug concentrations within hepatocytes and permitting the use of nucleoside analogues, the administration of which would otherwise be impossible due to extrahepatic toxicity.
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Article
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Article
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Chapter
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Chapter
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Chapter
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Chapter
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Chapter
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Article
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Chapter
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Chapter
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Chapter
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Chapter
The chapter discusses the persistence of foreign antigenic material in tissues, its chemical and biological properties, and speculations as to its role in immune mechanisms. A native antigen molecule soon changes after entering an intracellular environment. The persistence or retention of “antigen material” is important and not the original native substance that was used for immunization. Retention of antigen such as localization of antigen in normal and immune animals, retention of antigen in normal and immune animals, and retention of antigen in unresponsive animals is also provided in this chapter. It will also be seen that the retained antigen is not only a fragment of the immunizing material but that it is combined with normal tissue components such as ribonucleic acid (RNA) and under certain conditions with its specific antibody. Information related to the retention of foreign antigen material should provide a challenge to investigators of immune mechanisms. Future studies will probably revolve around investigations of chemically modified RNA in antibody-forming cells and studies of synthetic polypeptides or copolymers of various types with regard to their antigenic behavior. Characterization of retained antigen (physical properties and biological properties) and retained antigen and antibody formation is also discussed in this chapter.
Article
A sensitive biological test has been used to detect the persistence of minute traces of a foreign protein, bovine γ-globulin, in the blood and livers of rabbits intravenously injected with it, as an antigen. At various intervals after injecting these rabbits (donors) serum or liver tissue was transferred from them to the peritoneal cavities of normal or unilaterally adrenalectomized mice (recipients) with the aim of rendering the latter hypersensitive to the antigen that might be persisting in the transferred materials; a state of affairs detectable, 2 days later, by the appearance of signs of reversed passive anaphylaxis when the recipient mice were intravenously challenged with a strong anti-bovine γ-globulin rabbit serum. The protein persisted in the blood of the donor rabbits, in readily demonstrable amounts for 1 month, and in the blood of one animal, in minute traces, or as long as 6 weeks. It was detectable in the livers for 8 weeks. The persistence of bovine γ-globulin in rabbits, which form circulating antibodies to it well, is not as long as that in mice, which form antibodies to it poorly, since in previous work with the mouse the antigen was found (1) in the blood after 8 weeks and in the liver for 14 weeks. Nevertheless the antigen persists in the rabbit much longer than is generally supposed. Indeed it can be found in the liver all through the period in which circulating antibody is demonstrable in the blood. Explanations for the phenomenon have been suggested. Its significance in relation to the mechanisms of antibody formation is obvious.
Article
Viene descritta la particolare sensibilit dei macrofagi al coniugato amanitina-albumina. inoltre prospettata e discussa la possibile attivit antineoplastica di coniugati dell'albumina con sostanze inibenti la mitosi o la sintesi del DNA.
Article
Fluorescein-bovine albumin conjugates have been prepared and found not to differ appreciably in size, shape, and homogeneity from the precursor, bovine serum albumin. Fluorescein has also been conjugated to rat plasma proteins. Their disappearance rates from the circulation of rats correspond with those obtained from the use of isotope labeling. Their sites of localization in rat tissues were shown to be in the cytoplasm but not in the nuclei of Kupffer cells, fixed macrophages, granulocytes, and proximal renal tubules. Adsorption to endothelium was a characteristic finding. Extracellular localizations were predominantly in the lumina of blood vessels and proximal renal tubules (but never in the lumina of collecting tubules), and the interstitial fluid of skeletal and cardiac muscle (but not that of glandular organs such as the adrenals, liver, and spleen). BAC absorption from the skin of rabbits requires days whereas sodium fluorescein absorption is measured in hours, attesting to the persistence of the colloidal state of BAC in vivo. Fluorescein conjugates have been used to visualize the transcapillary passage of circulating proteins in the mesenteric circulation of frogs and rats by direct microscopic observation and found to diffuse slowly in the manner predicted for plasma proteins. The normal cutaneous vessels of the rat are impermeable in the gross to the labeled proteins; second degree burn promptly increases the permeability of these vessels rendering the presence of the label detectable in the gross in the skin. The process of labeling does not render guinea pig albumin antigenic, although slight antigenicity results from labeling whole plasma protein. It is believed that sufficient biological evidence is presented to support the conclusion that fluorescein-conjugated plasma proteins, particularly albumin, behave in vivo like their native precursors.
Article
Delivery of poorly soluble anticancer drugs can be achieved by employing polymeric drug delivery systems, capable of forming stable self-assembled nanocarriers with drug encapsulated within their hydrophobic cores. Computational investigations can aid the design of efficient drug-delivery platforms; however, simulations of nanocarrier self-assembly process are challenging due to high computational cost associated with the large system sizes (millions of atoms) and long timescales required for equilibration. In this work, we overcome his challenge by developing a multiscale computational approach in conjunction with experiments to analyze the role of the individual building blocks in the self-assembly of a highly tunable linear poly(ethyleneglycol)-b-dendritic oligo(cholic acid) block copolymer called telodendrimer. The multiscale approach involved developing a coarse grained description of the telodendrimer, performing simulations over several microseconds to capture the self-assembly process, followed by reverse mapping of the coarse grained system to atomistic representation for structural analysis. Overcoming the computational bottleneck allowed us to run multiple self-assembly simulations and determine average size, drug-telodendrimer micellar stoichiometry, optimal drug loading capacity, and atomistic details such hydrogen-bonding and solvent accessible area of the nanocarrier. Computed results are in agreement with the experimental data, highlighting the success of the multiscale approach developed here.
Article
SYNOPSISThe use of the direct fluorescent-labelling technique as a method for demonstrating the uptake of proteins by living cells is evaluated. This technique has been applied to the uptake of wheat-germ “lipase” and insulin by a selection of normal and malignant cell types. Some enzymatic activity of the “lipase”, which shows aliesterase activity, has been shown to remain after the labelling procedure. It was taken up in relatively large amounts by pinocytosis, and accumulated on the cell membrane of normal cells. Insulin did not appear to be taken up by pinocytosis.
Article
Lymphknoten und Knochenmark von Meerschweinchen wurden im Explantat auf verschiedene Weise mit Ziegenerythrocyten als Antigen zusammengebracht. In 3 von 49 Versuchen waren in vitro gebildete Hämolysine nachweisbar, darunter in einem Versuch in 2 Ansätzen. Vom Körper abgetrennte Zellen sind also zur Antikörperbildung fähig. Die Bedingungen, unter denen Antikörperbildung in vitro regelmäßig oder zumindest häufig erfolgt, haben wir nicht aufgefunden. Ein Züchtungsverfahren, bei dem wir Explantatzellen und Antigen in unmittelbaren Kontakt miteinander bringen konnten, führte nicht zu den erwarteten positiven Ergebnissen. — Möglicherweise entstehen Antikörper in vitro häufiger als sie nachweisbar sind. — Wir halten es für wahrscheinlich, daß der Vorgang der Antikörperbildung in vitro zwar ablaufenkann, daß dies aber tatsächlich nur selten geschieht, weil das Antigen in ungenügendem Maße oder zu spät von den Zellen in vitro aufgenommen wird.
Article
Improvements in a method for the specific microscopic localization of antigen in tissue cells are described. This method employs antibody labelled with fluorescein isocyanate as a histochemical stain, the specific antigen-antibody precipitate being made visible under the fluorescence microscope. Two isomeric series derived from nitrofluorescein are described.
Article
Cells from lymph nodes of rabbits injected repeatedly with bovine serum albumin were transferred subcutaneously to previously irradiated rabbits, and the recipients were immediately injected with bovine serum albumin. A good antibody response resulted. In a series of such animals killed on successive days, skin samples at sites of cell deposition were removed and examined by immunofluorescence and by light microscopy. In these tissues abundant plasmocytes were found to have multiplied and differentiated in a regular progression from immature, to medium, to mature plasmocytes. During the 6 days of the experiment the small plasmocytes accumulated until they reached 85 per cent of the total plasmocytic population. The mitotic index of the large and medium plasmocytes averaged 11 per cent, implying a generation time of 6.3 hours on the basis of a 1 hour mitotic time. This rate of growth is sufficiently rapid to account for all the plasmocytes on the 6th day as deriving from less than 1 per cent of the population initially transferred. This rate and the orderly progression in the evolution of the plasmocytic population, make it highly improbable that plasmocytes arise from transformation of lymphocytes, but rather indicate that they spring from specific precursors already present among the transferred cells.
Article
Following single intravenous injections of a foreign protein antigen, bovine gamma-globulin, into mice and rabbits, antigenic material persisting in the liver could be detected for several weeks. Ground liver tissue-taken from the mice and rabbits, just mentioned, either 4 or 6 weeks after injecting the antigen-when transferred repeatedly, at 2 or 3 day intervals, to the peritoneal cavities of normal, or unilaterally adrenalectomized, recipient mice, rendered the recipients sensitive to active anaphylaxis when they were challenged after a suitable interval by intravenous injections of the original antigen. The work throws some light on the state of the antigenic material that persists for 4 to 6 weeks in the livers of the donor animals. Obviously it is sufficiently unchanged, at least in its reactive groups, to engender in the recipient mice antibodies capable of reacting with the original antigen.
Chapter
The Polymorphonuclear LeucocyteCytoplasmic Granules of Polymorphonuclear LeucocytesMacrophagesThe Degradation of Bacteria Following Phagocytosis By Pmn Leucocytes and MacRophagesThe Fate of Antigens Within Phagocytic CellsSummaryReferencesDiscussion
Article
Normal and irradiated rabbits were given small doses of I131 bovine gamma globulin (BGG) and the rates of its loss from their blood were determined. The figures agreed with those of previous reports. 3 to 4 months later, both groups were reinjected with antigen. The control group gave an accelerated (anamnestic) rate of loss, indicative of the immune state. The irradiated group gave a response similar to that given by the control group on primary injection. Rabbits recover from 500 roentgens of x-ray in less than 2 months. Hence, rabbits given BGG soon after irradiation should become immune to this antigen in 2 months if they retain it. Since rabbits reinjected with BGG 3 to 4 months after irradiation did not give an accelerated response, the BGG must not have been retained over the period of time necessary for recovery; i.e., less than 2 months. As the rate of antigen loss is greater in normal than in irradiated animals, normal rabbits will have lost BGG as active antigen prior to the irradiated animals. The amounts of antigen used (0.25 to 0.4 mg./kg.) more nearly approximate the amounts present during disease than have the amounts used in previous studies of antigen retention. The hypothesis that protein antigens are lost as fast or faster than homologous proteins is discussed and the conclusion reached that this is a valid concept.
Article
1. Groups of normal rabbits were given, single intravenous injections of foreign proteins in doses of 1 gm. per kilo, bled at regular intervals for serologic studies, and sacrificed after varying lengths of time for pathological studies. The protein solutions used were of crystallized bovine serum albumin, bovine serum gamma globulin, and bovine serum. The experiments were planned, first, to correlate the sequence of pathological and immunological changes, and second, to compare the responses to two chemically and immunologically distinct plasma protein fractions and to the whole serum of the same species. 2. (a) The principal pathological lesions in rabbits given bovine serum were similar to those which have been previously observed following, the injection of horse serum and were characterized by widely dispersed but segmental acute inflammatory lesions of the arteries. These lesions were at their height 2 weeks after injection and showed marked repair at 4 weeks. (b) Crystallized bovine serum albumin produced lesions almost exclusively confined to the arteries which were at their height at 2 weeks, were healing at 3, and healed by 4 weeks. The lesions were less numerous and less intense than in animals given whole serum and were only found in some of the animals. (c) Bovine serum gamma globulin elicited quite different histologic sequences. The most striking lesions involved the glomeruli of the kidneys, and to a lesser degree, the heart. Lesions in the liver and joints were present but less conspicuous, and arterial lesions were rare and slight in degree. The lesions not only differed from those in rabbits given albumin in distribution but in timing, since they were most widespread and acute at 1 week and were healing at 2 weeks after injection. Moreover, lesions were observed in almost every animal. 3. Results of immunological studies were consistent with the interpretation that the pathological lesions were due to an antigen-antibody reaction in the tissues, as shown by the following: (a) Acute lesions were only observed when antigen was present and before antibody appeared in the circulation. (b) Healing of lesions was only observed (with one exception) when antigen had almost or completely disappeared from the circulation, usually with the appearance of antibody. (c) There was a correlation between the rapidity of evolution of the lesions and the rapidity with which the antigen disappeared from the circulation. (d) There was a rough correlation between the proportion of animals showing lesions and the proportion developing antibodies after the injection of a particular protein solution.
Article
Typhoid vaccine and sheep erythrocytes were injected subcutaneously into the feet of rabbits, and the subsequent formation of agglutinins and hemolysins in the popliteal lymph node was compared with the output of lymphocytes through the efferent lymph and with changes in the lymph node. Antibodies began to appear in the efferent lymph 2 to 4 days after the injection of the antigen and reached their highest titer after 6 days. This was preceded by a sharp rise in the output of lymphocytes through the efferent lymph, while in the lymph node there was lymphatic hyperplasia after preliminary infiltration of granulocytes and monocytes. This hyperplasia was first of a diffuse type, but was later superseded by large so called germinal centers, the latter lagging somewhat behind the rise in antibody titer. The fact that the tissue response accompanying the formation of antibodies was chiefly a lymphocytic one points to the lymphocyte as a factor in the formation of antibodies.
Article
1. The use of an antigen which can be seen within cells demonstrates that one may stimulate the phagocytic cells either of the liver and spleen or of the tissues and lymph nodes to produce antibodies. 2. The appearance of antibodies in the serum correlates with the time when the dye-protein is no longer visible within the cells and with the phenomenon of a partial shedding of their surface films. 3. It is thus inferred that the cells of the reticulo-endothelial system normally produce globulin and that antibody globulin represents the synthesis of a new kind of protein under the influence of an antigen. 4. An antigen is a substance which can specifically modify the synthesis of the cytoplasm of the cells of the reticulo-endothelial system.
Article
An antiviral principle is elaborated within the regional lymph nodes draining skin into which vaccinia is injected. The immunity conferred by clinical Jennerian vaccination may be largely of lymph node origin.
Article
1. A quantitative theory of the precipitin reaction based on the laws of classical chemistry has been tested on an azoprotein-antiprotein system and found to apply. 2. With its aid relationships may be deduced which permit the calculation of the behavior of an antidye serum over its entire range after a few quantitative chemical analyses have been made for antigen and antibody in the precipitate. 3. An empirical relation is also presented which further reduces the number of analyses necessary. 4. A study of supernatants in the inhibition zone has shown that the entire amount of dissolved antigen-antibody compound present is precipitated when supernatants are analyzed for antigen by the precipitin method.
Article
Agglutinins are formed within the draining lymph nodes of mice, following intradermal injections of killed cultures of microorganisms.
Article
1. Antisera to R-salt-azo-benzidine-azo-crystalline egg albumin give precipitates with crystalline egg albumin by virtue of their antidye content. 2. The quantitative course of the reactions with increasing amounts of antigen is very similar for the dye-antidye and egg albumin-anti-egg albumin systems, but differs markedly for the cross reaction between egg albumin and antidye. 3. A possible explanation for the occurrence of this one-sided crossreaction is given in terms of reactive groupings on the antigen and antibody. 4. A qualitative expression of the course of the cross-reaction is given in terms of the laws of classical chemistry.
Article
1. The preparation is described of a deep red protein dye, R-salt-azo-benzidine-azo-crystalline egg albumen, which contains no more than traces of protein with the original egg albumen specificity. 2. Based on previous publications of the writers, a quantitative method is given for the micro estimation of precipitin in the antisera to the dye. The method gives the actual weight of precipitin and may be applied to the determination of the maximum amount of precipitable antibody in any antiserum. 3. Data are given (1) on the influence of the period between the final injection and the bleeding on the precipitin content of rabbit antisera to the azo protein; (2) on the magnitude of the antibody response following the injection of multiple doses of the antigen varying within wide limits; (3) on the variations in the precipitin content of the sera of rabbits given successive courses of antigen injections; and (4) on the stability of antisera stored in the cold. 4. Four antisera were obtained in which over 100 times as much precipitin was recovered as the amount of antigen injected. This supplements the growing mass of evidence against the theory that specific antigen fragments are actually incorporated into the antibody molecule.
Article
Experiments are described which show that anaphylactic shock can be induced in animals sensitized with azoproteins by injecting them with azodyes containing the same azo components as the sensitizing antigen. The anaphylactic reactions are specific and occur with quantities of the dyes as small as fractions of milligrams.
Article
A standardized solution of a vital dye which escapes with some difficulty from the lymphatics of the ear of the mouse has been utilized in tests of the permeability of the lymphatic wall under various conditions. It has been found that this permeability is subject to great change. The slight pressure that suffices to prevent lymph flow from the ear,-an organ in which such flow goes on normally,-soon results in increased permeability of the obstructed lymphatics without as yet any perceptible dilatation of these vessels. Mechanical stimulation as for example a stroke with a blunt wire, or scratching so light as not to break the epidermis, results in a practically immediate, great increase in lymphatic permeability, which is sharply localized to the region pressed upon. This increase in permeability, though so great that even hemoglobin is let pass by the lymphatics, endures but a few hours. Warming the ear to 43 degrees C. or exposure to mild sunlight increases permeability considerably. Slight chemical irritation increases it greatly, though not so much that particulate matter is let pass. The edema developing as result of lymphatic obstruction or mechanical, thermal, or chemical stimulation is preceded by and associated with a large increase in lymphatic permeability. The facts are discussed in relation to their bearing upon fluid accumulation within the tissue. It is plain that influences within the realm of the normal suffice to increase lymphatic permeability and that those which lead to edema cause a very great increase in it. In proportion as this increase occurs the lymphatics cease to be channels demarcated by a semipermeable membrane. It seems certain that the changes must be in some part responsible for the local accumulation of fluid. There exist possibilities, on the other hand, of a correlation between the functionings of the blood and lymph vessels under certain pathological conditions, as during the resorption of edema.
Article
A technique has been developed for the demonstration of lymphatic capillaries in the ear of the mouse by means of vital dyes and for tests of their permeability under normal and pathological conditions. The lymphatics become visible as closed channels from which the dyes escape secondarily into the tissue. Some of them, cross-connections, with extremely narrow lumen, would seem ordinarily not to be utilized. There is active flow along the lymphatics of the mouse ear under ordinary circumstances. The movement of dye was always toward the main collecting system. The valves of the lymphatics as well as fluid flow prevented distal spread. There was in addition slow migration, apparently interstitial in character, but in the same general direction, of dots of color produced by the local injection of dye. The normal permeability of the lymphatics was studied with dyes of graded diffusibility. Their walls proved readily permeable for those highly diffusible pigments that the blood capillaries let through easily, but retained those that the latter retained. Finely particulate matter (India ink, "Hydrokollag"), they did not let pass. No gradient of permeability was observed to exist along them such as exists along the blood capillaries of certain organs. The observed phenomena of lymphatic permeability, like those of the permeability of the blood capillaries, can be explained on the assumption that the lymphatic wall behaves like a semipermeable membrane.
Article
From the experiments which have been reported, it follows that animals sensitized with one azoprotein react not only to the antigen used for the sensitization, but also to other azoproteins made up from the same simple azo-compounds and another protein. Although the specificity of the reaction has not yet been tested with various azo-components, its actual existence can reasonably be assumed on the basis of the phenomena observed in precipitation reactions. The sensitization is brought about with less facility than sensitization against the usual antigens and the effects are not uniform. Still, after sufficient treatment, 40 per cent of the animals succumbed with typical anaphylactic symptoms, mostly within a short time while 16 per cent showed severe symptoms. The experiments show that it is possible to make animals hypersensitive against a simple chemical group like para-arsanilic acid, and from this point of view connection would seem to be established with the phenomena of drug allergy in human beings. There is an essential difference, however, in that the sensitized animals did not react on injections of simple compounds such as para-amino-phenyl-arsanilic acid and phenyl-4-arsonic-acid-azo-tyrosine uncombined with protein. It remains to be determined whether under changed conditions positive results in this direction can be obtained. For this purpose it seems advisable to make experiments with isolated organs, according to the method of Schultz and Dale. While the simple substances failed to elicit direct reactions, they protected (as was foreseen by Doerr) against a subsequent injection of the active antigen. Similar compounds not containing the arsanilic acid group were considerably less active. The phenomenon is comparable to the inhibition of precipitin reactions already described. Considering the protection as a condition of antianaphylaxis, one would suppose that the simple substances mentioned are fixed by the cells in which the anaphylactic reaction takes place. It may be concluded that: 1. Animals can be sensitized through injections of one azoprotein-protein combined with diazotized para-arsanilic acid-against another compound containing the same azo-component but a different protein. 2. Injections of related simple compounds, as for instance para-arsanilic acid and phenyl-4-arsonic-acid-azo-tyrosine did not cause shock in the sensitized animals under the conditions of the present experiments. 3. The simple compounds mentioned and other related substances as well protect against the anaphylactic action of azoprotein, by inducing a state of antianaphylaxis.
Article
1. The injection of horse serum either in small or in large amounts in human beings is always followed sooner or later by the development of hypersensitiveness of the skin to subsequent injections of horse serum. For the development of this reaction serum disease is not essential. 2. The blood serum of most patients who suffer from an attack of serum disease following injections of horse serum shows anaphylactin and precipitin for horse serum. 3. Anaphylactin and precipitin cannot be demonstrated in the blood serum of patients treated with horse serum who do not later present symptoms of serum sickness. 4. The appearance of anaphylactin and precipitin precedes shortly recovery from the disease. 5. With the appearance in the serum of antibodies to horse serum in great concentration, the antigen rapidly diminishes or disappears. 6. It is probable that the extrusion of these antibodies into the circulation is the result and not the cause of serum sickness. Their presence serves to neutralize or destroy the antigen and thus determines the recovery from serum sickness.