Question

You have identified a monoclonal antibody (mAb) that effectively neutralizes one strain of Ebola virus (Zaire), but is less effective on another strain of Ebola virus (Sudan). Describe a process that you could use to modify the variable regions of the mAb heavy and light chains, with the hope to create a mAb that effectively neutralizes both virus strains.

Please explain the process step by step.

Answer 1

First, we need to identify a specific sequence of the variable region that recognize a common epitope in both viruses. To accomplish this, we need to use two groups of mice, one group will be in contact with Ebola virus (Zaire) and the other group will be in contact with Ebola virus (Sudan). After one or two weeks, the antibodies are extracted from the lymph nodes. Applying an Enzyme-Linked ImmunoSorbent Assay (ELISA) with a common antigen of both viruses (previously extracted and identified), you will determine which antibodies of both groups recognize the antigen. Once you obtained the specific antibody, you need to determine the amino acid sequence of the variable region (light and heavy chain) to introduce the specific DNA sequence in a mammalian expression system to synthetize the antibody. You need two plasmid vectors system, one for the light chain and other for the heavy chain which are going to be co-transfected into the mammalian expression system (antibody cell producer). After the co-transfection of plasmids, cell proliferation and protein synthesis, the cells are centrifuged and the protein is purified by affinity purification using the peptide tags chromatography columns. Finally, you can take a sample and use ELISA against the Fc region of the antibody to determine the concentration and presence of the new antibody.