The presence of alien histocompatibility antigens on the cell surface of the 3-methylcholanthrene-induced BALB/c (H-2d) fibrosarcoma C-1, was investigated by serological and transplantation studied. Absorption experiments with monospecific alloantisera showed that C-1 cells expressed their original private (H-2.4 and 31) and public (H-2.3, 8, 28, and 35) specificities. C-1 cells were also able to absorb monospecific antisera directed to the private specificity H-2.23 of the H-2k haplotype, as well as antisera to the public specificities H-2.1, 5, 11 11 and 25 (H-2k and in part H-2q, H-2a and H-2b haplotypes), which are absent from H-2d normal cells. Conversely, other alien specificities (H-2.2, 17, 30, 32, and 33) were not detected on C-1 cells. The C-1 cells were also unable to absorb the activity of an anti-Ia serum (1-28) directed to 1a.1, 2 and 19 (lak) specificities. Transplantation studies showed that resistance against the challenge of C-1 cells could be induced in syngeneic BALB/c mice by preimmunization with normal tissues from C3Hf and AKR (H-2k), A (H-2a) and C57BL/6J (H-2b) strains (expressing all or some of the extra H-2 antigens of the tumor) whereas no protection was obtained with DBA/2 (H-2d) or with W/Fu rat tissues. The anti-tumor activity could be passively transferred by BALB/c lymphoid cells immune to normal C3Hf, AKR, A, and NIH (H-2q) tissues, but no protection was achieved with lymphoid cells immune to DBA/2 or to W/Fu normal rat tissues. These data indicate that foreign H-2 antigens are expressed on C-1 tumor and that they might function as tumor-associated transplantation antigen which was shown to be present and individually distinct on this sarcoma by appropriate in vivo tests.

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W e have previously shown that the methylcholanthrene-induced BALB/c fibrosarcoma C-1 syngeneically transplanted in vivo expressed, i n addition t o i t s original H-Zd and tumor-associated transplantation antigens, also the foreign H-2Kk 23, 1,5, 11.25 specificities. In the present study we have investigated the expression and the immunogenicity of these antigens on an in vitro line of C-1 tumor. The binding of C57BL/6J o r C3Hf anti-BALB/c sera o r that of monospecific alloantisera t o H-2d specificities on plated C-1 cells (evaluated by the indirect isotopic '251-antiglobulin assay) showed that the expression of H-Zd antigens was very low o r undetectable on the tumor kept in vitro. Absorption by the in vivo and in vitro maintained C-I cells of the complement-dependent cytotoxicity on BALB/c normal lymphoid cells of the C75BL/6J anti-BALB/c serum confirmed the lower expression of H-2d antigens on C-1 kept in vitro as compared t o the in vivo transplanted tumor. The H-Zk antigens found on C-1 were also examined by the above methods using BALB/c anti-C3Hf, anti-AKR, anti-C57BL/6J polyspecific sera and also monospecific alloantisera directed t o H-Zk antigens. The C-1 cultured cells displayed a significant reduction of H-Zk antigens as compared t o the in vivo C-1 tumor, the antigen 5 being undetectable on the in vitro cells.Allogeneic and syngeneic sera raised against in vitro o r in vivo C-1 cells were tested on either BALB/c o r C3Hf normal lymph-node cells by the complement-dependent cytotoxicity assay. A significantly higher titer of cytotoxic antibodies t o H-Zd and t o H-2k was obtained i n the allogeneic and syngeneic sera respectively, after immunization with the in vivo C-1 cells as compared t o sera raised against the in vitro tumor line. Transplantation studies showed that both in vitro and in vivo C-1 lines possess the same tumor-associated transplantation antigen whose expression was nonsignificantly reduced on the cultured cells.

SUMMARY By immunizing BALB/c (H‐2d mice against normal tissues from C57BL/6J (H‐2b), C3Hf (H‐2k) and DBA/2 (H‐2d), but not from AKR (H‐2k) strains, resistance was induced to the subsequent challenge of the ‘syngeneic’ methyl‐cholanthrene‐induced BALB/c sarcoma ST5; lymph node cells from allo‐immune BALB/c mice were also able to exert a parallel cytotoxic effect against in vitro cultured ST5 cells. The involvement of foreign H‐2 specificities in the observed cross‐reactions was ruled out by absorptions of H‐2 monospecific sera and by the interallelic combinations used, thus suggesting that non‐H‐2 histocompatibility antigens were responsible for the above findings. By using the indirect isotopic antiglobulin assay, BALB/c anti‐C57BL/6J and anti‐C3Hf polyspecific sera were found to bind specifically to cultured ST5 cells. C57BL/6J and C3Hf, but not DBA/2, lymph node cells were able to absorb the anti‐ST5 activity of the anti‐C57BL/6J serum. These results indicated that ST5 cells expressed on their surface at least two different sets of foreign non‐H‐2 antigens: one shared by C57BL/6J and C3Hf tissues, and detected by both cell‐mediated and serological techniques; the other one belonging to DBA/2 tissues, and revealed mainly at the cell‐mediated level.

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