The Core offers types of hybridoma fusion methodologies: PEG-assisted fusion (the standard methods for chemically induced fusion of cells) and Electrofusion.(using short pulses of an electric field to induce fusion of cells) to form hybridomas.
Regardless of the method employed, antigen-reactive B cells are obtained from immunized animals and fused with myeloma partner cells to create a hybrid cell called a hybridoma, which secrete a specific monoclonal antibody. All hybridomas are born with an inherent instability resulting from having a double chromosome number after formation. The tend to lose some of their chromosomes as the hybridoma divides. If the lost chromosome encodes for the immunoglobulin gene, the hybridoma will no longer be capable of secreting an antibody. We refer to this hybridoma as being unstable.
Subcloning is used to identify unstable hybridomas, as well as insure a hybridoma is truly monoclonal. It is strongly recommended that subcloning be repeated at least twice or until every subclone exhibits positive response to the antigen. Negative subclones indicate that the hybridoma is either a mixture of cells secreting the desired mAb and cells that are not OR the hybridoma is unstable because many of the cells are losing ability to secrete mAb.