THE USE OF PRECIPITIN ANALYSIS IN AGAR FOR THE STUDY OF HUMAN STREPTOCOCCAL INFECTIONS
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Abstract:
It has been shown by agar precipitin tests (Ouchterlony and Oakley) that human sera may contain from 0 to 5 antibodies against antigens present in a partially purified streptolysin O preparation, and from 0 to 7 antibodies against antigens in a crude ammonium sulfate concentrate of the streptococcal culture supernate used. These antigens were prepared from a Group A hemolytic streptococcus (strain C203S). Strong evidence was presented suggesting that some of the bands seen with streptolysin O concentrate represented antibody reponses to streptococcal antigens heretofore undescribed. Tests were also carried out with other streptococcal antigens, including streptokinase-desoxyribonuclease mixture from Group C streptococci (varidase-Lederle), crystalline proteinase, proteinase precursor, C carbohydrate, and sonic vibrated streptococcal cell extracts (group A, C203S). Fewer bands were seen with these preparations, and with some they were quite uncommon. The observations indicated that the predominating antibody responses in human streptococcal infections were to extracellular products of the micro-organisms, and only very slightly and infrequently to intracellular antigens. The human sera studied included sera from patients with active or convalescent rheumatic fever, and non-rheumatic subjects suffering from a variety of illnesses. As was expected, the rheumatic subjects showed antibody responses to many more of the antigens present in these preparations than did the nonrheumatic group. Pooled normal human gamma globulin was found to contain many of the antibodies found in potent human sera. This finding confirmed the antigen-antibody nature of the bands seen with individual sera. The epidemiological significance of these findings with gamma, globulin was briefly discussed. It was found that rabbit, guinea pig, and human antibody precipitin bands join quite readily in the Ouchterlony tests. This finding adds another tool for the identification of the precipitin bands found with human sera. Evidence was obtained which indicated differing immunological specificities of two samples of streptococcal desoxyribonuclease, one from Group A, the other from a Group C streptococcus. The value of these technics as representing a new approach to the study of human infectious disease was discussed.Keywords:
Precipitin
Streptolysin
Streptococcus Pyogenes
Ouchterlony double immunodiffusion
Group A
Summary Seven highly purified antigens of known molecular weights ranging from 30,000 to 2,000,000 were studied by the Ouchterlony gel diffusion technique. It was found that the curvature of the precipitin line is related to the molecular weight of the antigen, i.e., a straight line is obtained when the antigen has approximately the same molecular weight as the antibody; if the line curves away from the antibody reservoir, the antigen is heavier than antibody; if it curves towards the antibody reservoir, the antigen is lighter than antibody. The theoretical and practical aspects of these findings are discussed.
Precipitin
Ouchterlony double immunodiffusion
Immunodiffusion
Molecular mass
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The effect of date extract on growth and hemolytic activity of S. pyogenes was examined. It was found that 5% DE caused 78 % growth inhibition. However, 20% DE inhibited the growth by 86%. 5% DE inhibited hemolysin and streptolysin O activities by 43% and 24% respectively,while 20% caused 95 and 91 %inhibition.
Streptolysin
Streptococcus Pyogenes
Hemolysin
Growth inhibition
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Summary A theoretical discussion of the precipitin patterns obtained with cross-reacting systems, as studied by the Ouchterlony gel diffusion technique, is presented. The conditions that result in single or double spurs are described, and experimental examples are given.
Ouchterlony double immunodiffusion
Precipitin
Agar gel
Double diffusion
Immunodiffusion
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Citations (23)
A theoretical discussion of the precipitin patterns obtained with cross-reacting systems, as studied by the Ouchterlony gel diffusion technique, is presented. The conditions that result in single or double spurs are described, and experimental examples are given.
Ouchterlony double immunodiffusion
Precipitin
Agar gel
Immunodiffusion
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Citations (26)
Cereolysin, streptolysin O, and perfringolysin O formed precipitin lines that completely fused when reacted with horse antitetanolysin by Ouchterlony immunodiffusion and formed precipitin lines that showed either partial or complete fusion when diffused against horse antistreptolysin O or antiperfringolysin O.
Precipitin
Ouchterlony double immunodiffusion
Immunodiffusion
Streptolysin
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Citations (27)
The Ouchterlony technique, single radial immunodiffusion and fluoro-electro-immunodiffusion (FEID) for the detection of α-fetoprotein (AFP) had their own merits. FEID was at least 30 times as sensitive as the Ouchterlony technique for the detection of serum AFP. Its great sensitivity elicited a positive reaction in sera which were previously negative by the Ouchterlony technique.
Ouchterlony double immunodiffusion
Precipitin
Immunodiffusion
Radial immunodiffusion
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Streptolysin
Streptococcus Pyogenes
Strain (injury)
Group A
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Precipitin
Ouchterlony double immunodiffusion
Immunodiffusion
Immunoprecipitation
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Streptolysin
Streptococcus Pyogenes
Hemolysin
Cytolysin
Exotoxin
Group A
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The successful classification of Group A streptococci by the capillary precipitin technique requires a complete series of M type antisera which are sufficiently potent and specific to give unequivocal type-specific reactions with all the serotypes. Specific antisera for this purpose have been prepared by absorption with heterologous streptococci. Unabsorbed antisera have been employed here in the Ouchterlony double-diffusion agar-gel test to identify the M type of streptococci. Techniques have been developed for making this method of M typing fully reliable. The results reported here confirm and amplify the original findings of Michael and Massell (3). With crude HCl extracts and unabsorbed M type antisera, a precipitin line due to the M protein and another to the group-specific carbohydrate are the two major reactions observed. These reactions, however, are usually readily distinguishable. There was a surprising lack of cross-reactive precipitin lines due to non-type-specific protein antigens in the extracts. Although many of the unabsorbed M type antisera can be employed in the double-diffusion tests, the group-specific antibody must be removed from some of the unabsorbed antisera to avoid confusing cross-reactions. Absorption of these antibodies has been achieved by means of a specific immunoabsorbent column prepared from para-aminophenyl-beta-N-acetylglucosamine and cyanogen bromide-activated Sepharose. Excellent agreement was observed between the M typing results obtained on 117 field strains by the conventional capillary precipitin method and the Ouchterlony double-diffusion method.
Ouchterlony double immunodiffusion
Precipitin
Immunodiffusion
Antigenicity
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