| When antibodies are derived from a direct | | | | cells. Culture may be in vitro, or in vivo via |
| immune response they are heterogeneous. This | | | | injection into the mouse peritoneal cavity. The |
| means that B-lymphocytes derived from common | | | | cells are harvested from the ascitic fluid resulting |
| stem cells differentiate into many different cell | | | | from this. |
| lines, each producing immunoglobulins to a | | | | Monoclonal antibodies are widely used as primary |
| particular epitope on the immunogen. The serum | | | | immunoglobulins where specificity to a single |
| from an immunised animal will contain numerous | | | | epitope is required over an extended period of |
| epitope-specific antibody clones (hence it is | | | | time. Following culture, they are characterised to |
| polyclonal) which may comprise several Ig classes | | | | assess their titre, their class and subclass. Ab titre |
| and subclasses. Conjugated polyclonal antibodies | | | | is the dilution factor necessary to get a signal in a |
| make useful secondary immunoglobulins in IHC | | | | given immunoassay, for example Western blot or |
| assays. | | | | ELISA (enzyme-linked immunosorbent assay). |
| A cell line derived from a specific B-cell clone | | | | Because each Ig class and subclass has specific |
| produces monoclonal antibodies, of which there | | | | characteristics, class determination is important to |
| are thousands on our antibody database. | | | | determine purification and modification procedures. |
| However, B-cells isolated from a heterogeneous | | | | For example, protein A is widely used for affinity |
| serum population cannot be cultured in useful | | | | purification of mouse IgG. However, it has a low |
| amounts. Thus we use hybridoma technology, the | | | | affinity for the subclass IgG1, and so purification |
| established method of producing monoclonal | | | | of this subclass should be carried out using protein |
| immunoglobulins since 1975. Isolated | | | | G. |
| B-lymphocytes from immunised mice are fused | | | | We at Novus Biologicals have a wide range of |
| with murine myeloma cells. This creates immortal | | | | isotyping and purification protocols, giving us a full |
| monoclonal hybrid cell lines specific to known | | | | and comprehensive antibody database covering all |
| epitopes. The hybridomas can be stored, and then | | | | the latest advances in protein biology. |
| recultured to keep producing more monoclonal | | | | |