Question

How would a monoclonal antibody to a viral protein be prepared?

Answer 1

Ans: Monoclonal antibodies are made from the hyrbidization of myeloma B-cells (carcinogenic B-cells) and spleen cells. The hybrids are selected through HAT medium. I order to make a monoclonal antibody for a viral protein; the viral protein is administred inside the host (mostly mice) and spleen cells against this viral protein is screened using ELISA method. These spleen cells are then mixed with Myeloma cells which are carcinogenic B-cells which does not produce HGPRT enzyme which is an enzyme essential for the salvage pathway of nucleotide synthesis. These mixture are plated on HAT media which is a selection media. HAT media contains Hypoxanthine, aminopterin and thymidine; Since myeloma cells does not contains HGPRT enzyme so they cannot utilize hypoxanthine for nucleotide synthesis through salvage pathway also exposing cells to aminopterin makes them unable to use de novo pathway of nucleotide synthesis. So myeloma calls can neither use salvage pathway nor de novo pathway, so they can’t able to survive in HAT media.Unfused spleen cells cannot grow indefinitely because of their limited life span. Only fused hybrid cells, referred to as hybridomas, are able to grow indefinitely in the medium because the spleen cell partner supplies HGPRT and the myeloma partner has traits that make it immortal (similar to a cancer cell). These survived hybrid cells positively selected and further grown to obtain monoclonal antibody which has one and only one target, that viral protein. So in this way monoclonal antibody is prepared for the viral protein.