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Effect of "anti-NK" sera on NK and ADCC activities was examined to study further the relationship between the effector cells for NK and ADCC in specific pathogen-free miniature swine. An appropriate dilution of "anti-NK" sera that had no effect on ADCC activity not only inhibited NK activity but also eliminated NK cells in the presence of C which supports the presence of NK cells distinct from K cells for ADCC.

A cytotoxic monoclonal antibody, SFR8-B6, produced by immunizing KGH rats with partially purified glycoprotein from JY, a human B lymphoblastoid cell line, detects HLA-Bw6, a unique public antigen on the HLA-B molecule. By cytotoxicity, SFR8-B6 has complete concordance with the presence of Bw6 as defined by alloantisera, and its reactivity segregates with HLA in families. Because sera detecting Bw4 and Bw6 are rare, this monoclonal antibody has value as a perpetual standard. In addition, SFR8-B6 offers potential as a probe for investigating the molecular relationships of HLA antigens.

We have developed a monoclonal antibody with HLA-DR5 serologic specificity. The antibody, SFR3-DR5, binds specifically to DR5-positive lymphoblastoid cell lines, and immunoprecipitates alpha- and beta-chains characteristic of DR antigens from them. Cytotoxic activity of the antibody segregates with the DR5-bearing haplotype in a family. The antibody reacted with the cells of 16 of 17 DR5 individuals and was negative on all DR5-negative cells tested. SFR3-DR5 reacted weakly with PWM-activated cells of the single DR5 individual whose B lymphocytes were unreactive with the monoclonal antibody by cytotoxicity. Possible interpretations of these results are discussed.

During the course of screening new T-H-2 region congenic strains of mice constructed from the C57BL/6 and B6-H-2k strains, a new cell surface polymorphism, designated dtc-1, was identified by cell-mediated lympholysis techniques. The dtc-1 antigen can be found on both Con A- and LPS-stimulated lymphoblasts, peritoneal macrophages, and SV40-transformed mouse embryo fibroblasts. Lysis of dtc-1+ targets by CTL is H-2Dk restricted. All inbred strains tested are dtc-1+, with the exception of the B6-H-2k strain, which is dtc-1-, and several congenic strains directly derived from B6-H-2k. Because B6/Boy and AKR/Boy, the parents of the B6-H-2k strain, are dtc-1+, the dtc-1- phenotype may be the result of mutation in the locus specifying the cell surface molecule that carries this antigen. Segregation analysis of the dtc-1+/dtc-1- polymorphism demonstrated that this locus is not linked to T or H-2. The dtc-1 antigen thus identifies yet another cell surface polymorphism and adds another immunologically defined genetic marker to the murine genome. Furthermore, the dtc-1 system indicates the need for reevaluation and restandardization of congenic strains of mice derived from the B6-H-2k congenic strain.

An anti-HL-A 3 antiserum with cross-reacting activity for HL-A 1 and HL-A 11 was subjected to isoelectric focusing over a pH 5-8 gradient. The cytotoxic activity of the serum focused into three distinct peaks, one at the basic end of the gradient, one between pH 6 and pH 7 and one at the acidic end. The first and second peaks reacted with HL-A 3 positive cells and HL-A 3 negative cells positive for cross-reacting antigens. The third peak reacted only with HL-A 3 positive cells.

The role of HLA antigens in the generation of cytotoxic cells in CML has been investigated. Cytotoxic effector cells were generated in MLC among HLA-A or HLA-A and HLA-B disparate, HLA-D identical siblings, and among HLA-A and HLA-B disparate, MLC identical (%RR less than or equal to 2 3.6) unrelated individual. The data indicate that HLA-D differences and poliferative MLC responses as measured by 3H-thymidine incorporation are not requisite for the in vitro generation of cytotoxic cells and suggest the existence of a CML-S locus (loci) distinct from HLA-A, HLA-B and HLA-D. The degree of cytotoxicity generated in a proliferative versus a "nonproliferative" MLC was comparable. In addition, these studies demonstrate that antigens other than the currently definable HLA-A, HLA-B, HLA-C, and HLA-D can serve as target determinants in cell-mediated lympholysis.

It has been determined that alpha-2 macroglobulin (alpha 2M) is more suppressive of a mixed lymphocyte response (MLR) when complexed with proteinase than in its "native" state. Other alpha 2M preparations showed a moderate level of MLR suppression, but it is unlikely that this is a result of interaction with cellular proteinases. A panel of other proteinase inhibitors (alpha 1 PI, SBTI, BPTI, TLCK) did not suppress the MLR to the same extent as alpha 2M either when bound with or free from trypsin. A dose-responsive pattern of MLR suppression similar to that observed with purified proteinase-complexed alpha 2M was seen with serum containing proteinase-complexed alpha 2M. The population of cells that apparently conveys the suppressive property is the adherent cells (putative monocytes), which can reduce the MLR almost as well as unfractionated cells when exposed to alpha 2M. Most of these properties of alpha 2M were demonstrable in "serumless" medium with qualitative similarity to the MLR obtained in cultures performed with conventional serum supplemented medium. It was found that alpha 2M-trypsin complexes must be presented at or near culture initiation and remain in contact with the cells for a minimum of approximately 4 hr to have its optimum effect.

The evolution of HLA specificities has been toward ever-increasing refinement; one example is a subdivision of HLA-B5, the supertype specificity originally defined as 4a. HLA-B5 can now be further subdivided into Bw51 amd Bw52 by serologic means. Whereas the specificity Bw51 can be detected by specific sera, the identification of Bw52 must frequently be deduced from knowledge of B5 and Bw51, although serology has progressed rapidly. There has been no comparable development in identifying fine specificities by cellular cytotoxicity in populations. We have now found that cytotoxic effectors of exquisite specificity can be generated against Bw52 by sensitization of cells from a Bw51 donor and vice versa; Bw51 and Bw52 can in this way be recognized with equal ease. This may set a precedent for recognizing fine specificities of other HLA antigens that cannot yet be identified serologically or can be identified only imprecisely. These fine distinctions may have great relevance in allotransplantation and in understanding disease susceptibility.

Serum antibodies to adenovirus (ADN), cytomegalovirus (CMV), herpes simplex virus (HSV), influenza (INF), para-influenza (PAR), mumps (MUM), coxsackie B4 (Cox B4) and B5 (Cox B5) viruses were measured from 584 individuals belonging to 21 Indiana Amish families. Sex and age effects on antibody responses to cytomegalovirus were observed. Age effect on CMV, HSV, INF, PAR, MUM responses were also found. The percentage of responders to some of the viruses was shown to be age dependent, but the levels of antibody response were not affected by the difference in age. A familial basis for the antibody response was demonstrated. Attempts at demonstrating association between HLA haplotypes and responses were not successful. The unlikelihood of predominantly HLA-associated control of viral antibody response was discussed.

Natural killing (NK) capacity was evaluated in peripheral blood mononuclear cells from 14 patients with well defined primary immunodeficiency disorders and compared with the activity of those cells in antibody-dependent cell-mediated cytotoxicity (ADCC) assays against antibody-coated erythrocyte (killed primarily by monocytes) and lymphoid or tumor targets (killed exclusively by lymphoid cells). A selective inability to lyse antibody-coated lymphocyte targets was observed with cells from patients with x-linked agammaglobulinemia, suggesting the involvement of either a different lymphocyte subpopulation or membrane receptor for NK and ADCC, or that a different functional susceptibility exists for the two types of killing. The only immunodeficiency state in which lymphocyte NK activity was found to be lacking was severe combined immunodediciency disease.

H-2 antigens from EL4 (H-2b) and RLmale 1 (H-2d) leukemias were solubilized with deoxycholate and partially purified based on their adherence to a Lens culinaris hemagglutinin column. In double reciprocal experiments we have shown that these preparations specifically inhibit conjugate formation between target cells and alloimmune peritoneal lymphocytes, a preparation rich in cytotoxic T lymphocytes.