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The immunologic response of neonatal and of older rabbits to the tissue (transplantation) antigens of pooled rabbit leucocytes was studied, the test system being the suppression of formation of agglutinin to Shigella paradysenteriae by transferred lymph node cells which had been incubated with Shigella antigen. In the active induction of the suppressive effect on the transferred cells it was found that neonatal rabbits reacted as vigorously as 1 kg rabbits to the prior injection of a given number of rabbit leucocytes pooled from prospective donors of lymph node cells. The suppressive effect was dose-related, and within the range of number of leucocytes used was similar for both age groups of recipients. An attempt to detect a difference in the response during the first few days after leucocyte injection, before the full suppressive effect is reached, failed to show any difference between rabbits of the two age groups. Since it had been found possible to transfer the suppressive effect passively with sera obtained from older rabbits injected with rabbit leucocytes, attempts were made to do so with sera obtained from neonatal rabbits injected with similar numbers of pooled adult rabbit leucocytes. No consistent suppression of transferred lymph node cells was observed with sera from neonatal rabbits, even with relatively large amounts of such serum. In sera of rabbits which had been injected with rabbit leucocytes at the age of 4 to 6 weeks, suppressive antibody could be detected. When anti-rabbit-leucocyte serum obtained in adult rabbits was injected into neonatal and 1 kg recipients at a given volume per gram of animal weight the suppressive effect of the serum was of similar extent in the two groups of recipients. In the adoptive transfer of the lymph node cell suppressive effect, by cells of lymph nodes draining the sites of injection of pooled rabbit leucocytes, it was found that the popliteal lymph node cells of neonatal rabbits were as effective as those of 1 kg rabbits. Splenic cells of neonatal rabbits were also effective, when an adequate number of rabbit leucocytes had been injected intravenously. Thus, in conferring adoptive immunologic response, as in active immunization, the neonatal rabbits were as effective as the older rabbits in their response to homotransplantation antigens, in contrast to the considerable difference in concentration of the suppressive antibody in sera of neonatal and older rabbits injected with rabbit leucocytes.

A fraction of rabbit lymph node cells and splenic cells has been prepared by differential centrifugation of sonically disrupted cells a highspeed centrifugal sediment from the supernate of a lower speed centrifugation. This cell fraction gives complement fixation with rabbit antirabbitleucocyte serum and gives evidence of containing nearly all the complementfixing isoantigen of such cells.

In earlier studies it was shown that if rabbit lymph node cells were incubated with Shigella-trypsin filtrate and transferred to other, recipient, rabbits, agglutinins to Shigella appeared subsequently in the sera of the recipients. However, if leucocytes from the donor rabbits were injected into the recipients at a suitable interval prior to lymph node cell transfer, agglutinins to Shigella did not appear after cell transfer. In the present study, a number of experiments were done which bear on the immunologic character of this suppressive effect on the transferred cells, the "pre-injection effect." X-irradiation of the recipients before the injection of donor leucocytes led to a decrease in the extent of the pre-injection effect, i.e. to the appearance of higher agglutinin liters, in the recipient animals. It was found possible to transfer the pre-injection effect by cells of rabbit lymph nodes draining sites of injection of rabbit leucocytes. No such effect was obtained with heated aliquots of these cell suspensions or with lymph node cells from sheep-erythrocyte-injected donors. It was also possible to transfer the pre-injection effect passively by serum. Suppression of transferred lymph node cells was observed regularly after injection of serum pooled from groups of rabbits which had been injected with leucocytes pooled from the blood of 60 to 70 rabbits. The active material in anti-leucocyte serum could be precipitated in the globulin fraction of the serum and could be removed by absorption with rabbit lymph node cells. In the course of experiments on the passive transfer of the pre-injection effect by antisera of individual rabbits to leucocytes of individual donor rabbits, evidence was obtained of the existence in the sera of some normal rabbits of antibodies to some of the rabbit leucocyte antigens.

Shigella paradysenteriae organisms were incubated in a solution of trypsin and then removed from the suspension by Seitz filtration. Serologic tests with anti whole Shigella serum indicated that the filtrate contained antigenic material derived from the organisms. When lymph node cells from rabbits not previously injected with Shigella were incubated in vitro with such filtrates and then transferred to irradiated recipients, agglutinins to Shigella appeared in the sera of the latter. The transfer of heated suspensions of cells was not followed by the appearance of agglutinins in irradiated recipient rabbits. Non-irradiated recipients of lymph node cells incubated with the filtrate also developed agglutinin titers, to a lower level than irradiated animals. Agglutinins did not appear in the sera of the majority of non-irradiated recipients of heated cells. The addition of excess antiserum to either the filtrate or the cell suspension before the incubation of these two materials resulted in a marked reduction in the subsequent agglutinin titers of recipient animals. However, if antiserum was added at the conclusion of the usual 30 minute incubation of cells and filtrate, or even after 5 minutes of incubation, there was no reduction in the agglutinin titers of the recipients. Cytologic examination of aliquots of a number of the suspensions of cells prepared for transfer revealed that approximately 98 to 99 per cent of the cells belonged to the lymphocytic series.

A study of the effect of alloantibody-containing ascitic fluid globulin of mice on the retention of skin allografts has been carried out using, as a tentative measure of the relevant antibody, the titer of the antibody in such globulins which can suppress the production of hemolytic antibody plaques by donor strain spleen cells. When globulins with sufficiently high suppressive titer were available in sufficient quantities, pools of such globulins were injected into normal mice which were given skin allografts in the same strain combination as that used for production of the alloantibodies. Among globulins, all of which were above a given threshold of suppressive titer and all within a given range, four kinds of effect were found on the time of retention of the allograft: accelerated rejection in almost all of the grafted mice (relative to control mice injected with normal ascitic globulin), or accelerated rejection in some of the grafted mice, or no apparent effect, or, finally, prolonged retention of the graft. There was no relation between the suppressive titer of the globulin pool and its effect on the time of rejection of the skin graft. The fact that anti-H-2 antibody-containing globulin pools could cause, in given instances, these opposite effects of accelerated rejection and prolonged retention of the allogeneic skin grafts suggested the participation of at least two alloantibodies with the same specificity but with differing biological effect, the fate of the graft depending on the relative concentrations of the two. The possible roles of two such competing antibodies are being explored.

Antibody-producing cells have been identified among cells obtained from efferent lymphatic vessels, the thoracic duct, and peripheral blood. These cells, which produced plaques of hemolysis and which were quite rare (20 to 50 per million), due in most instances to 19S antibody, were located and studied by electron microscopy. Of the antibody-producing cells found in these three sites there were several features common to all: small size (5 to 8 µ), generally spherical shape, approximately central position of the nucleus, and retention in the nucleus of the condensations of chromatin characteristic of the lymphocyte. The differences among the cells of these sources were largely in the relative amount and state of organization of the organelles of the cytoplasm. In cells of the efferent lymphatic vessel and the thoracic duct, the endoplasmic reticulum showed a range from relative scarcity to considerable numbers of well organized channels. Between these extremes were cells with a considerable amount of endoplasmic reticulum, the channels disorganized and sectioned at various angles. The cytoplasmic matrix of all of these contained a profusion of polyribosomes. Antibody-producing cells obtained from peripheral blood showed, around the roughly spherical nucleus, a ring of cytoplasm which was narrow, but wholly organized into parallel lamellae of endoplasmic reticulum, with many polyribosomes between these, and a large Golgi body. Some similarities and some differences of these cells, in comparison with antibody-producing cells obtained from lymph nodes, have been indicated.

Efferent lymph of the popliteal lymph nodes of rabbits was collected 4 days after a single footpad injection of SRBC. Thin-layer agar plating was done to isolate plaque-forming cells of the lymph for electron microscope examination, and the numbers of plaque-forming cells (PFC) in cells from the lymph and lymph nodes were determined. Of 71 PFC of lymph isolated and examined, 93% were lymphocytes, most of them with signs of substantial levels of physiologic activity. The cytoplasm showed an abundance of free ribosomes and many finger-like projections. The endoplasmic reticulum (ER) was barely detectable in most of the active lymphocytic PFC, and in some, a few short narrow channels of ER could be seen. Approximately one-fifth of the lymphocytic PFC presented an appearance of senescence, with signs of degeneration: rounded cells, with amorphous nuclear chromatin, and very few microvilli. The remaining 7% of the PFC of the lymph showed an unusual combination of features: small round cells with a narrow ring of cytoplasm which, however, contained well-organized channels of ER. Such cells had been found only among PFC of peripheral blood of the rabbit. The number of PFC per million cells was higher in the lymph than in the suspensions of lymph node cells. In both the contralateral lymph node and its efferent lymph, the number of PFC was less than 1% that of the injected side.

Cells were teased from the popliteal lymph nodes of rabbits which had not been injected with dysentery bacilli. These cells were incubated with dysentery bacilli in vitroand transferred to X-irradiated recipients. Agglutinins were detected in the sera of such recipients on the fourth day after transfer. When the cells were heated prior to transfer agglutinins did not appear during the first week after transfer. Similar results were obtained when suspensions of lymph node cells and suspensions of dysentery organisms were injected separately into irradiated rabbits.