Hybridoma technology is a powerful technique used to produce monoclonal antibodies (mAbs), which are highly specific proteins capable of recognizing and binding to specific antigens.
The technique involves the fusion of antibody-producing B cells with immortalized myeloma cells, creating hybridoma cells that continuously produce large amounts of a single type of antibody.
Production of Monoclonal Antibodies
1) Immunization
An animal (usually a mouse) is immunized with the antigen of interest.
This stimulates the production of specific antibodies by B cells within the animal's immune system.
2) Harvesting of B Cells
B cells producing the desired antibodies are harvested from the animal’s spleen or lymph nodes.
3) Fusion of B Cells with Myeloma Cells
The harvested B cells are fused with immortalized myeloma cells using polyethylene glycol (PEG).
The resulting hybridoma cells combine antibody specificity with the immortal growth properties of myeloma cells.
4) Selection and Cloning of Hybridoma Cells
Hybridoma cells are screened (e.g., ELISA, flow cytometry) to identify those producing the desired antibody.
The selected hybridoma cells are cloned to establish stable cell lines capable of mass-producing monoclonal antibodies.
Purification of Monoclonal Antibodies
Once the hybridoma cells are successfully cultured, the monoclonal antibody must be purified from the cell culture or animal tissue.
The purification process involves the following steps:
Step 1: Harvesting of Cells
The hybridoma cells are collected from the culture medium or tissue using centrifugation or filtration.
Step 2: Cell Lysis
The collected cells are lysed (broken open) to release the monoclonal antibody using sonication or homogenization.
Step 3: Purification
The antibody is purified from the lysate using techniques such as:
Column chromatography
Protein A/G affinity chromatography (which specifically binds antibodies, separating them from other proteins).
Applications of Hybridoma Technology
1) Diagnostics
Monoclonal antibodies are integral in diagnostic tests detecting specific antigens or antibodies.
Example: ELISA (enzyme-linked immunosorbent assay) diagnostic tests.
2) Therapeutics
Monoclonal antibodies serve as targeted therapeutic agents in treating diseases such as cancers and autoimmune disorders.
Examples
Infliximab (Remicade): Treats autoimmune diseases (e.g., Crohn’s disease, rheumatoid arthritis).
Herceptin: Treats breast cancer.
Rituximab (Rituxan): Treats non-Hodgkin’s lymphoma.
3) Research
Monoclonal antibodies facilitate the study of protein and cell functions.
Used in laboratory experiments for purification or isolation of specific molecules.
Examples of Products Developed Using Hybridoma Technology
Infliximab (Remicade): Therapeutic monoclonal antibody.
Herceptin: Therapeutic antibody targeting breast cancer.
Rituximab (Rituxan): Therapeutic antibody for lymphoma treatment.
ELISA Diagnostic Tests: Common diagnostic laboratory tests.
