Ascites is the term that describes the fluid that fills the peritoneal cavity due to inflammation. For in vivo monoclonal antibody production, hybridoma cells are injected into the peritoneal cavity to resulting in ascites fluid containing the desired antibody.
The ascites method is one of the quickest for producing large amounts of monoclonal antibodies. One mouse can produce 3-5 ml of ascites fluid after undergoing several extraction cycles after inoculation. Inoculating multiple mice simultaneously can produce large batches of a monoclonal antibody.
Ascites fluid can contain antibodies at concentrations that range from 1-3mg/ml. This range is higher than what is secreted by hybridoma cells that do not express well in an in-vitro system. However, since ascites also contain antibodies that were produced in the mouse that bind other antigens, the titer for each batch of ascites should be determined.
Ascites antibodies are useful for the same assays as antibodies produced through animal-free methods. Routine applications include western blotting, immunohistochemistry, immunocytochemistry, and ELISA. Since ascites contain proteases and would degrade; purification is highly recommended before proceeding with any assay. In addition, because ascites antibodies are a mixture of the desired antibodies and nonspecific antibodies from the host, low specificity may be an issue. To improve signaling, we recommend extra blocking and washes steps and incubating the sample at 4°C for 12-16 hours.
The disadvantage of making antibodies through the ascites is that it requires animals. The mice need daily observation and must be kept in a clean facility otherwise they risk contamination from other biological agents such as viruses, bacteria, or mycoplasma.
