Marilyn L. Bach

Differential function of major histocompatibility complex antigens in t-lymphocyte activation.

The antigenic systems of the major histocompatibility complex can be subdivided into those which are serologically detectable and those which are detected in tests with mixed lymphocytes. The two systems have different roles in the activation of separate populations of T lymphocytes.

Histocompatibility Matching Vi. Miniaturization Of The Mixed Leukocyte Culture Test: A Preliminary Report

A miniaturized method for the mixed leukocyte culture (MLC) test is described which results in substantial savings in cells, medium, and time. The test is done in microtiter plates using 0.2-ml culture volumes. Concentrations of responding and stimulating cells are varied, depending on the experiment. Significant discrimination between isogeneic and allogeneic mixtures is possible when cultures are labeled for a 16–20-hr period starting at 48 hr. The method measures MLC identity, as previously defined, and quantitates in a manner significantly correlated with the macro method.

Cell Mediated Immunity: Separation of Cells Involved in Recognitive and Destructive Phases

The mixed leukocyte culture (MLC) and the cell mediated lympholysis (CML) assays are used as in vitro models of the afferent, or recognitive, and efferent, or destructive, phases of the homograft reaction. Activity in both of these tests has been related to differences at the major histocompatibility complex, HL-A in man and H-2 in mouse. Recent evidence suggests that the presumed cell surface differences which lead to cell proliferation in MLC are different from those which act as a target for CML. Data are presented providing further support for this hypothesis; in addition separate cell populations may respond to the differences which activate cells in MLC and to the differences which serve as targets for CML. There thus appears to be a dichotomy both for genetic control of, and cell populations involved in, the recognitive and destructive phases of cell mediated immunity.

Genetic and Immunological Complexity of Major Histocompatibility Regions

There are genetic differences within the major histocompatibility complex of the mouse which lead to skin graft rejection but which cannot be detected serologically. When confronted with these differences on allogeneic cells, lymphocytes proliferate in vitro. In other cases, in vitro lymphocyte proliferation but no skin graft rejection is associated with loci that are linked to but genetically separable from the loci controlling the serologically defined antigens.

Antigenic requirements for triggering of cytotoxic t lymphocytes.

The cotnplexity of T lymphocyte subclasses that respond to alloantigens can be analyzed from at least two perspectives: first, by identification and separation of the different subclasses on the basis of their adherence properties, differentiation antigens present on their surface or other characteristics; and second, by analyzing the alloantigenic systems to which the cells respond. We have focused primarily on the second of these approaches utilizing genetically-defined antigens of the major histocompatibility complex (MHC) as the stimulating factors evoking T lymphocyte responses. It is from this perspective that we shall analyze the requirements for the triggering of T lymphocytes, discussing the cytotoxic T lymphocyte (CTL) in detail with attendant comments about helper and suppressor T lymphocytes. We have used a number of different test systems that appear to provide information in this regard; four of these will be discussed. First, the in vitro generation of helper, suppressor and cytotoxic T lymphocytes in a primary mixed lymphocyte culture (MLC) sensitization system; second, secondary

Leukemia-Associated Antigens in the Mixed Leukocyte Culture Test

Patients with acute leukemia (blast cells in the peripheral blood) manifest antigens in the one-way mixed leukocyte culture test. Leukocytes from normal patients and leukocytes of patients in remission do not clearly show these antigens. These antigens could be leukemia-associated antigens in man.

β2-Microglobulin: Association with Lymphocyte Receptors

β2-Microglobulin (β2m) is a low-molecular-weight protein constituent of lymphocyte membranes. Amino acid sequence analysis has revealed a high degree of homology between the β2m and certain regions of immunoglobulin molecules, suggesting a possible recognition function for the β2m, in analogy with the immunoglobulins. The data presented demonstrate that highly specific antiserum against β2m blocks lymphocyte reactivity against allogeneic cells in mixed leukcocyte cultures and against phytohemagglutinin, both of which processes presumably function via a cell surface receptor on thymus-derived (T) lymphocytes. There is very little inhibition of T lymphocyte rosette formation with sheep red blood cells. The findings suggest a possible relation between the β2m and recognition units on the T lymphocyte surface.

Chloral Hydrate: A Solvent for Biological Membranes

A buffer system conitainin chloral hydrate, taurine, and bromopyridinium lactate was used to dissolve several biological membranes and separate their protein components by polyacrylamide gel electrophoresis. This solvent system was capable of separating molecules of similar size on the hasis of their charge and allows easy recovery of the proteins Thus. aqueous chloral hydrate is an effective solvent for biological membranes.

Cartilage-hair hypoplasia: Effect of thymus transplants☆☆☆

A patient with cartilage-hair hypoplasia manifesting deficient cell-mediated immunity (CMI) but normal antibody-mediated immunity (AMI) suffered from a progressive vaccinia reaction. Neutralizing virus antibodies were demonstrated but no in vitro reactivity to vaccinia antigen nor to allogeneic cells or phytohemagglutinin could be demonstrated. Three thymus transplants were accomplished over a period of 4 months. Biopsy revealed growth and enlargement of one of the transplants. Increased numbers of small lymphocytes appeared but the total number remains diminished. The child has recovered from the infection but remains deficient in cell-mediated immunity. Lymphopenia persists and only modest responses to phytohemagglutinin and allogeneic cells can be demonstrated at this time. This experience suggests that isolated CMI deficiency will not routinely respond to thymus transplants in contrast to the experience reported in patients with the DiGeorge syndrome.

Lymphocyte reactivity in vitro. VII. The effect of polymorphonuclear leukocytes on lymphocyte response.

SUMMARY One-way mixed leukocyte culture (MLC) tests were performed using, in all cases, mitomycin C-treated stimulating cells from healthy university students. Responding cells from four groups were tested: hospitalized patients with malignancy, age- and sex-matched hospitalized patients without known malignancy, age- and sex-matched nonhospitalized controls, and students who were younger than the persons in the other three groups. Whereas approximately 30% of the variation in response was unaccounted for by known variables, the great majority of the remaining variation could be accounted for by the number of polymorphonuclear leukocytes present in the culture. These results suggest that quantitation of MLC tests may be better after removal of polymorphonuclear leukocytes from the culture—a step we have now instituted in our laboratory.