C. L. Oakley

The Relative Transmission of the Fractions of Papain Hydrolyzed Homologous

Molecules of rabbit γ-globulin are split into three nearly equal pieces by papain. Trace labelled with 131I these three fractions are transmitted by the yolk-sac to the circulation of the 24-day rabbit foetus in significantly different quantities. Fraction III is transmitted nearly as readily as whole γ-globulin, whereas fractions II and I are transmitted only 1/5 and 1/10 as readily, respectively, as fraction III. It is suggested that fraction III, which contains most of the antigenic groups of the original molecules, also has the configuration recognized by the cells as homologous γ-globulin.

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During the investigation of a number of epidemics of Clostridium welchii food poisoning (Hobbs, Smith, Oakley, Warrack & Cruickshank, 1953) the opportunity arose of examining some 500 different strains of Cl. welchii for toxigenicity. Work on so large a scale made it essential to develop a routinefor this purpose, and after several trials we devised the folio whig sequence of tests, which have proved entirely satisfactory in our hands, and may be of interest to others.

-globulin from the Uterine Cavity to the Foetal Circulation in the Rabbit

Antibodies pass from the uterine lumen of pregnant rabbits into the foetal body fluids. It has been shown that antibodies formed in rabbits enter the foetal circulation by this route much more readily than those formed in horses or cows. It is the intention of this paper to investigate further this selective admission to the foetal circulation, to determine if there is a differential selection of heterologous antibodies according to the species in which they were produced and if this selectivity changes with foetal age. Antitoxins were employed throughout and were prepared in rabbit, man, guinea-pig, dog, horse and cow, against diphtheria or tetanus toxin or Clostridium welchii α-toxin. Mixtures of sera from two or three of these species, each serum containing a different antitoxin, were administered so far as possible. The antitoxin concentrations observed in the experimental samples were corrected to unit concentration in the serum administered, the term concentration quotient (c.Q.) being used for this quantity. Measured in this way the entry of antitoxin to the foetal serum varied with the species in which the antitoxin was produced; rabbit, man, guinea-pig, dog, horse and cow forming a descending series in that order. The entry of antitoxin into the amniotic fluid, or the foetal stomach contents derived from it, does not vary with the species in which the antitoxin was produced. This is so also for the maternal serum. Thus the species in which an antitoxin is produced affects the concentration at which it will appear in the foetal serum but not that at which it will appear in the other foetal body fluids or in the maternal serum. The entry of antitoxin prepared in rabbits to the foetal circulation increases with foetal age from 20 to 26 days after copulation and decreases thereafter.

Routine Typing of Clostridium welchii.

SUMMARY: Three antigenic components (α, β, γ) are present in toxic culture-filtrates of Clostridium histolyticum. Of these α is the lethal and necrotizing toxin, β is a collagenase, and γ is a cysteine-activated proteinase which attacks altered collagen (e.g. hide-powder or azocoll) but not native collagen. The β and γ-enzymes both attack gelatin. By methods based on the properties of these antigens, Cl. histolyticum antisera can be tested for the corresponding antibodies.

Selection of antitoxins by the foetal membranes of rabbits.

Natural and pepsin-digested rabbit antitoxins were compared with respect to their transmission to the foetal circulation in rabbits. The experiments were so designed that the foetuses were exposed simultaneously to the two antitoxins in rabbit serum injected into the uterine cavity. The natural antitoxin was transmitted readily, but the pepsin-digested antitoxin was not transmitted at the lowest concentrations tested, so that concentration quotients for pepsin-digested antitoxin ranged from < 1/6 to < 1/30 of those for natural antitoxin. It is concluded that the pepsinized moiety of an homologous antitoxin molecule is transmitted less readily than the whole.

The alpha, beta and gamma antigens of Clostridium histolyticum (Weinberg & Séguin, 1916).

Antibodies can be detected in the amniotic fluid and in stomach contents of rabbit foetuses exposed to immune serum following injection into the uterine lumen or into the maternal circulation. It is shown that the positive reactions obtained with foetal stomach contents were due to the presence of specific antibodies derived from the immune sera to which the foetuses were exposed. By means of injections of immune rabbit and bovine sera into the embryonic membranes of 24-day rabbit foetuses, it was demonstrated that the antibodies in the stomach contents are derived from the amniotic fluid and not from the foetal circulation. There is a pronounced tendency for antibodies to be concentrated in the stomach, concentrations of 32 and 64 times the titre of the amniotic fluid being observed. Although antibodies of rabbit origin readily pass through the yolk-sac splanchnopleur into the foetal circulation, they do not do so through the stomach wall in detectable amounts.

The relative transmission of natural and pepsin-refined homologous antitoxin from the uterine cavity to the foetal circulation in the rabbit.

Globulins originating in another species are treated in the circulation of the rabbit in the same way as is homologous globulin. Twenty-four hours after intravenous injection of mixed antitoxin preparations the relative concentrations of rabbit and bovine antitoxins in the foetal sera are unchanged. There is no evidence of differential treatment of homologous and heterologous protein after entry to the foetal circulation. Since serum globulins taken up from the uterine cavity of the rabbit by the yolk-sac splanchnopleur reach differential concentrations in the foetal serum depending upon the species of origin of the globulin, selection must occur during passage of the protein through the tissues of the yolk-sac splanehnopleur itself. The rate of rem oval of globulin from the foetal circulation during the first 24 hours is not more rapid than in the adult. No evidence could be found of any passage of in tact globulin across the placenta from the foetal to the maternal circulation.

The accumulation of antibodies in the stomach contents of foetal rabbits.

SUMMARY: Evidence is provided in support of the view of MacLennan, Mandl & Howes (1958) that some strains of Clostridium histolyticum produce a cysteine-inactivated proteinase—the δ-antigen.

Protein Selection in the Yolk-Sac Splanchnopleur of the Rabbit: The Fate of Globulin Injected into the Foetal Circulation

Antitoxins were employed to measure the relative rates of entry of antibodies into the foetal circulation, and into the amniotic fluid, from the uterine lumen in rabbits on the 24th day of gestation. Antitoxic sera prepared in rabbits, cattle and horses against diphtheria toxin, tetanus toxin and Clostridium welchii α-toxin were used, since tests for antitoxins are both sensitive and quantitative. These sera, each immune to one antigen, were mixed and the mixture was injected into the uterine lumen. Analysis of the relative concentrations attained in the foetal sera and amniotic fluids showed that whereas rabbit antitoxins entered the foetal blood at a rate at least fifty times greater than bovine or equine antitoxins, all three antitoxins entered the amniotic fluid at almost identical rates. The concentrations attained in the substance of the membranes traversed provide evidence suggesting the operation of at least two distinct mechanisms. The entry of rabbit antibodies into the foetal circulation may depend on an active process of absorption and secretion by the cells. The entry of both homologous and heterologous antibodies into the amniotic fluid and of heterologous antibodies into the foetal blood, on the other hand, may depend on a process of ‘seepage’ between the cells.