Hen's egg yolk contains at least two antigen-binding subclasses of IgG, derived from the hen serum and transmitted t o the chick. IgM and IeA. absent in volkInstitute of Animal Physiology, Babraham, and in newly hatched chick serum, were detected in the white of unembrvonated Cambridge+

No abstract

The large amounts of IgA made by plasma cells in the gut and mesenteric nodes produce high levels of IgA in the intestinal and thoracic duct lymph which are difficult to reconcile with the low levels found in the blood of most species.In rats, blood serum levels of 0.05-0.18 mg/ml have been reported while mesenteric or thoracic duct lymph with about one-third of the total immunoglobulin level of blood contain 0.6 mg/ml (1-5). Assuming a flow rate of 1-3 ml/ h for thoracic duct lymph of rats and a blood plasma vol of 8 ml the amount of IgA which daily enters the blood from the thoracic duct is 20-50 times greater than the circulating plasma pool. The evident disappearance of this IgA from the blood suggests that it is either catabolized Very fast or exported. The availability of IgA from rat plasmacytomas (6) now makes it possible to purify enough IgA to explore these possibilities, Preliminary experiments had shown (a) that IgA disappeared from the blood very much faster than similarly labeled IgG2; (b) that feces collected during the day of injection contained antigenically intact radiolabeled IgA; and (c) that in accord with Lemaitre-Coelho et al. (7) rat bile had IgA levels some 10 times higher, and IgG levels 30-50 times lower, than those of blood serum. As the liver of healthy mammals contains no cells considered capable of immunoglobulin synthesis, it seemed likely that the biliary IgA was derived from the blood by active transport. Materials and MethodsImmunoglobulins and Antisera. IgA was isolated from the ascitic fluid of Lou/Wsl rats bearing the IR461 plasmacytoma. After salt precipitation the material was fractionated twice on a 2.6 × 100 cm column of Ultrogel Ac22. The IgA formed by IR461 is heterogeneous with regard to size, and, like normal rat IgA, mainly oligomeric (5). The fraction least contaminated with other proteins (Fig. 1) had a sedimentation rate of 13.2 S; it contained a-macroglobulin detectable in immunoelectrophoresis and estimated as 0.3 mg/ml. Other contaminants account for less than 5% of the total protein (5.4 mg/ml) so the fraction was about 90% IgA.IgG2 was made from serum of Lou/Wsl rats bearing the IR33 plasmacytoma which secretes IgG2a. The material used in the clearance studies, eluted from DEAE cellulose with 0.01 M phosphate buffer pH 7.4, contained some IgG2b but no IgG1.Antisera obtained by immunizing rabbits with purified myeloma IgA contained anti-idiotypic and other unwanted antibodies, most of which were absorbed out with a pseudoglobulin

Lymph-borne immunoblasts were labeled in vitro with 125I[]iodo-deoxy-uridine, washed and returned by intravenous injection to the sheep from which they had been collected. Twenty h later the sheep were killed and the distribution of the immunoblasts was determined by assaying the radioactivity in various organs. Immunoblasts from the efferent lymph of peripheral somatic lymph nodes (PSLN) went mainly to the spleen, lungs and other PSLN, while immunoblasts from intestinal lymph went mainly to the small gut. This ability of intestinal immunoblasts to home to the gut was demonstrated also in the sterile environment of fetuses in utero; apparently the migratory behavior of immunoblasts, like that of small lymphocytes, is not primarily "antigen-driven". A technique was devised for the collection of peripheral (i.e. afferent to the mesenteric node) intestinal lymph which was found to contain 10-20 times the numbers of small lymphocytes that occur in the peripheral lymph from other tissues. Immunoblasts from peripheral intestinal lymph also homed to the gut. The immunoglobulin content of immunoblasts was studied by making detergent extracts of lymph cells, by applying immuno-peroxidase techniques to cell films and by investigating the incorporation of 14C-labeled amino acids into immunoglobulins by immunoblasts in vitro. Immunoblasts from both somatic and intestinal lymph contained and made IgG and IgM, but many intestinal immunoblasts contained and made IgA. It is not known whether this immunoglobulin mediates the extravasation of immunoblasts into the gut. Nonetheless, there is compelling evidence that there are two major migratory pathways for lymphoid cells; one through the gut-associated lymphoid tissue and the other through the somatic-splenic lymphoid tissues.

The rapid and active transport of polymeric IgA from the blood to the bile (1, 2) is selective (3, 4), which suggests the existence of a specific receptor for this molecule. The liver cells responsible for the transfer have been identified as the hepatocytes by autoradiography and electron microscopy of the livers of rats killed 5, 30, and 60 min after the injection of radiolabeled IgA (5). 5 rain after being injected, the radiolabeled IgA appeared to be bound to the sinusoidal membrane of the hepatocytes. We have found that intravenously injected radiolabeled IgM, IgG1, and monomeric IgA did not appear in the bile at all, and that the transport into bile of s-IgA (i.e., polymeric IgA bound to secretory component [SC]) was both considerably slower and quantitatively less than that of polymeric IgA devoid of SC. Studies of the transport into the bile of IgG2a, IgE, IgM, monomeric and polymeric IgA, and s-IgA with isolated perfused rat livers gave similar results: only polymeric IgA that lacked SC was actively transported (6). These facts suggest that SC may be the receptor for IgA on the surface of rat hepatocytes as it is on the epithelial cells from human gut (7). The availability of hepatocytes in short-term monolayer culture (8), together with that of specific antibodies to rat SC, has enabled us to test this hypothesis by experiments in vitro. Materials and MethodsPreparation of lmmunoglobulins, Free SC, and Specifically Purified Antibody to SC. The preparation and properties of the polymeric rat IgA formed by the IR 461 plasmacytoma have been described (1). Rat IgG was the fraction of normal rat serum eluted from DEAE cellulose with 0.01 M phosphate buffer at pH 7.2. Normal rat bile is a good source of both s-IgA and free SC (9). Pooled bile was first treated with 15-20 mg/ml of eholestyramine resin (Cuemid; Merck, Sharpe and Dohme, Ltd., Hoddesdon, Herts., U. K.), to reduce the amount of pigment. After 10-fold concentration by ultrafiltration, followed by centrifugation, the bile was fractionated on DEAE cellulose. All protein, reactive with antibody to either a-chain or to SC, eluted together with a phosphate buffer (0.05 M, pH 6.2) that contained 0.2-0.3 M NaCI. Gel filtration of this material on a column of Ultrogel AcA34 (LKB Instruments, Croydon, Surrey, U. K.) gave an excluded peak that contained IgA and a second peak that contained free SC and albumin. Further gel filtration of these two fractions on Ultrogel AcA22 and AcA34, respectively, yielded a preparation of pure polymeric s-IgA and one of free SC that still contained some serum albumin; the antigenic properties of the biliary SC are shown in Fig. I a. Rabbit antiserum obtained after three injections of a crude SC preparation was absorbed first on an immunoadsorbent column of normal rat serum proteins coupled to AH-Sepharose (Pharmaeia Fine Chemicals, Inc., Piscataway, N. J.) and then on a similar column of IR 461 myeloma IgA. The specificity of the anti-SC serum is shown in Fig. 1 b. Specifically purified antibody was made from this antiserum...