Polyreactive antibodies are natural antibodies capable of binding multiple antigens and are commonly found in normal and diseased individuals. To explore mechanisms involved in polyreactivity, and study other biological properties of polyreactive antibodies we performed in vivo and in vitro studies with human monoclonal and polyclonal antibodies. We found that polyreactive antibodies are able to bind multiple antigens simultaneously. The carbohydrate moieties of immunoglobulins are not associated with polyreactivity. The half-life of polyreactive Abs is significantly shorter compared to its monoreactive counterpart and is isotype independent. Using biotinylated antigens as probes to detect antigen-reactive B cells, we found that the cell populations positively selected have a higher percentage of polyreactive antibody-producing B cells than negatively selected populations, and the difference between them is about 10 fold. Thus, antigen binding might be used as a functional marker for isolating polyreactive antibody- producing B cells. Several studies have shown that the majority of polyreactive antibodies are produced by the CD5 antigen-bearing B cells. To understand the relationship between the expression of the CD5 gene and the production of polyreactive antibodies, we cloned, mapped and determined the exon-intron boundaries of the genomic form of the gene. Two targeting vectors have been engineered to mutate the CD5 gene either at the second or second and third exons in ES cells. The totipotency and ability of ES cells to contribute to the development of the germ line of the animal has been tested by making chimera mice.