Given the importance of membrane protein targets (multi-spanner), private and academic laboratories have tried numerous approaches to raise antibodies against them.
The most common way of generating these antibodies is immunization with whole cells over-expressing the target protein. This allows membrane protein targets to be displayed in their native conformation without mechanical or detergent disruption. Typically, stable murine cells expressing a human membrane protein are used to immunize mice. Attempts of using transiently-transfected cells, membrane preparations, and cells with the highest expression levels have been reported to improve this approach, with varying degrees of success. However, for many important membrane protein targets the ability of cell-based immunogens to elicit high-quality antibodies has been limited by low membrane-protein expression, the abundance of non-specific proteins, and target protein toxicity during cell line selection.
Another classic approach is to design peptide antigens derived from the extracellular loops for antibody production. Unfortunately, these peptide antigens could not fold in correct tertiary structures as within multi-spanner, thus they are not suitable for making conformational antibody against multi-spanners.
The solve these problems, we developed MPATTM platform (MPAT=Membrane Protein Antibody Technology). We applied the most recent methods of de novo design and structural biology to create a new platform to construct multi-spanner (G-protein-coupled receptor (GPCR) or ion channel) mimic protein antigen (MMPA). MMPA not only maintains structural characteristics of the whole multi-spanner extracellular domain, but also makes it feasible to express in whole water-soluble form with high purity. The revolutionary technology allows us to generate antibodies against GPCR or (or ion channel) much more efficiently.