Generation and Characterization of Hybridomas from HUG Transgenic JH Knockout Mice and JH Knockout Mice.

Abstract

Dr. Pollock?s lab studies transgenic mice that express a mouse/human chimeric antibodies (immunoglobulin) heavy chain gene called HUG. We want to determine the effect of the HUG heavy chain on expression of the mouse?s endogenous immunoglobulin (Ig) genes. From the results of previous studies we hypothesized that the HUG does not up-regulate the initiation of ? rearrangement. However, it has been difficult to determine the relationship between the HUG and the expression of ? because it was possible that the endogenous mouse heavy chain influenced the production of ? in the HUG+ mice. To eliminate the rearrangement of endogenous heavy chain in the HUG+ mice they were bred to JH knockout mice (JH-/- mice). The resulting HUG+ JH-/- mice allow us to study the effect of HUG transgene on ? expression. Our objective of this summer was to fuse splenocytes from HUG+ JH-/- and JH-/- mice with NSO cells in order to immortalize HUG+ JH-/- and JH-/- B cells. We tested the hybridomas for both secreted and cytoplasmic HUG and ? by ELISA. There was a lower percentage of HUG+/?+ cells found among HUG+ JH-/- hybridomas compared to HUG+ hybridomas. All JH-/- hybridomas were HUG-/?-. From these results, we conclude that the expression of ? may not be related to spontaneous rearrangement of light chain, but related to the HUG transgene. In the future experiments, we are planning to sub-clone our hybridomas in order to determine the actual number of hybridomas that are positive for HUG and/or ?.