The investigator will study the process of antigen selection and hypermatutation, and how they contribute to B cell memory, in the mouse response to phenylarsonate (Ars). This project involves the identification of clones in the Ars response at different stages of their differentiation, and the development of a correlation between this maturation stage and position in the spleen relative to the putative sites of antigen selection and somatic mutation. The Ars response is a well-characterized system by many laboratories. Two broad aims are proposed that are subdivided into several subaims. First, the investigator will characterize the anti-Ars response of A/J mice via the determination of spatial organization of anti-Ars-specific clones with the location of antibody forming cells (AFC) and germinal centers (GC). The first subaim will address the appearance and timing of Ars-specific clones by the use of an anti-Ars V gene antibody and PCR analysis. The investigator will distinguish between two principle models for antigen-driven B cell maturation. A second subaim will employ the same methodologies to evaluate the spatial organization of memory anti-Ars clones. The second broad aim will utilize the investigator s transgene system for study. In the previous period, the investigator has developed a transgene model where an anti-Ars specific rearranged IgVh gene is able to be recruited into the anti-Ars response (and undergo somatic mutation). Using these transgenic mice, they will study the organization of the transgene-encoded anti-Ars clones and B cell maturation. In the first subaim of this section, the investigator will compare the transgene-encoded clonotype (high affinity to Ars) with endogenously emerging clones (low affinity to Ars) for their population of spatial sites of B cell maturation, AFC and GC. For this study an additional antibody reagent will be employed that is more specific for the transgene clonotype. In the second subaim of this section, using the same general methodologies, they will characterize the relationship between affinity for Ars by these two antibody types and the somatic mutation levels of these genes. In a third subaim they will determine whether there is a positive relationship between sites of apoptosis in a GC and the relative incidence of affinity maturation.
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