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Culture supernatant fluids from spleen cells from C57BL/10 or BALB/c mice neonatally treated with semiallogeneic (B 10.D2 x B10)F1 cells to induce haplotype-specific suppressor T cells and restimulated with macrophages syngeneic at I-A with the allogeneic haplotype encountered as neonates contain a soluble factor capable of suppressing primary in vitro antibody responses of normal syngeneic spleen cells in a non-antigen-specific manner. This haplotype-specific suppressor factor, TsF-H, has also been recovered in culture fluids of a T cell hybridoma produced by fusion of the AKR thymoma BW5147 and the haplotype-specific suppressor T cells. TsF-H is inactivated by low pH (3.5) trypsin, for 30 min at 50 degrees C, and has a molecular weight in the range of 45,000 to 68,000. Studies with specific immunoabsorbents demonstrate the presence of determinants encoded by the I-A subregion of the haplotype of the T cell producing TsF-H but not I-J subregion or immunoglobulin constant-region determinants on the TsF-H. Suppression is restricted to primary in vitro antibody responses, and not secondary antibody, mixed lymphocyte, or cytotoxic lymphocyte responses by spleen cells syngeneic at the I-A subregion of H-2 with the T cell producing the factor. The properties and activities of TsF-H and the haplotype-specific suppressor T cell are compared and contrasted with antigen-specific and genetically restricted suppressor T cells and their factors.

Immunization of goats and mules with human thrombin resulted in an antiserum that reacted only weakly with the parent molecule, prothrombin. Some of the antibodies in this antiserum showed a greater affinity for thrombin complexed to its naturally occurring inhibitor, antithrombin-III, than for active thrombin. An antiserum against the human thrombin inhibitor, antithrombin-III, produced 2 precipitin lines against human serum but only 1 against plasma. The 2nd line in serum was shown to represent precipitation of a complex of thrombin with antithrombin-III. The neoantigens appearing in antithrombin-III after complex formation were also present in complexes prepared with purified clotting factor Xa and antithrombin-III. Since purified host (mule) thrombin was also capable of causing formation of the neoantigenic sites when complexed to human antithrombin-III, it seems likely that these determinants result from interaction in the host between the immunogens (either human thrombin or antithrombin) and the appropriate interacting host protein (mule antithrombin-III or thrombin, respectively). Studies by radioimmunoassay showed that the antibodies formed are not completely specific for the neoantigens since they also react to a lesser extent with the free proteins.

An E. coli tRNAfMet fragment [C-A-U-A-A-C-C-C-G-A-A-G-G-U-C-G-U-C-G-G (bases 35-f54)] containing the anticodon triplet has been synthesized by the phosphotriester method involving protected oligonucleotide blocks. Di- or tri-nucleotide blocks were prepared by condensation of 2'-O-(o-nitrobenzyl) nucleotide derivatives and used for the synthesis of pentanucleotide blocks. The 5'-hydroxy, heterocyclic amino and internucleotide linkage were protected with monomethoxytrityl, acyl and p-chlorophenyl groups, respectively. The 3'-phosphates of the pentanucleotides, except for the GUCGG block where 2'-O-benzoyl 3'-O-(o-nitrobenzyl) N-isobutyrylguanosine was used, were protected with p-chlorophenyl and anilido groups. The anilido groups were removed by treatment with isoamyl nitrite and the 3'-phosphodiesters of resulting pentamers were activated with mesitylenesulfonyl nitrotriazolide to give protected decanucleotides in yields of 61-89%. The two decanucleotides were condensed similarly to yield the protected eicosanucleotide in a yield of 59%. The product was deblocked and purified by ion-exchange chromatography on DEAE-Sephadex A-25 and characterized by enzymatic hydrolysis after labelling the 5'-end by phosphorylation using polynucleotide kinase and [gamma-32P]ATP.

Droplet counter-current chromatography (DCC) a novel liquid-liquid partition technique, is a useful method for the separation of glycolipids. After a series of investigations on suitable experimental conditions for DDC, mouse brain chloroform-methanol-0.5% CaCl2-n-propanol (50:60:40:6) by the ascending method with 500 standard columns. GM1 and cerebroside sulfuric ester (CSE) from whale brain glycolipid fraction were separated by DCC into two peaks and three peaks, respectively, with chloroform-methanol-water-n-propanol (50:60:40:2) and 500 standard columns. The occurrence of two peaks of GM1 was due to the differences of sphingenine and fatty acid compositions. In the former peak of GM1, the majority of long chain base and fatty acid were 18:1 sphingenine and stearic acid, respectively, while the latter contained 18:1 (32%), 20:1 (68%) sphingenine and 16:0 (14.5%), 18:0 (61.4%), and 20:0 (21.0%) fatty acids. The three peaks of CSE were due to the difference of carbon chain length and to the presence or absence of a hydroxyl group in fatty acids. The first polar fraction (CSE-I) had hydroxy 23:0 (34%) and 24.1 (53.4%) fatty acids and the second fraction (CSE-II) contained hydroxy 24:0 (70.5%) and 25:1 (12.8%) fatty acids, while the third fraction (CSE-III) had normal 23:0 (11.9%), 24:1 (56.4%), hydroxy 25:0 (11.4%) and 26:1 (9.7%) fatty acids. Preparative DCC was also applied to the glycolipid fraction from Tay-Sachs' brain to give 242 mg of practically pure GM2 from 700 mg of crude brain extract without any preliminary purification.