Repetitive Immunization Multiple Site (RIMMS) Introduction for Hybridoma Generation

A new immunization technique was reported by Kilpatrick and Bynum. Based on protein immunization, DNA immunization, and cell immunization, combined with the repetitive immunization multiple sites immunization (RIMMS), the preparation time of monoclonal antibodies was greatly shortened. They successfully produced monoclonal antibodies with high affinity against the immunogen by fusing draining lymph node cells of mice immunized with antigen or DNA for 7-13 d with tumor cells stably transfected with the Bcl22 gene.

Technical Principle for Antibody Generation by RIMMS

The maturation of T cell-dependent B-cell responses is associated with the formation of germinal centers in the dynamic microenvironment that develops in secondary lymphoid tissues. Dendritic cells take up and deliver antigen from antigen presentation sites, eliciting an immediate T cell-dependent B-cell response in regional lymph nodes.

3–5 days after immunization, antigen-sensitized T and B cells interact with APC in a discriminative manner, leading to the formation of germinal centers. All the components of antigen-specific immunoglobulins develop in the germinal center as B cells expand and undergo antigen-driven maturation and selection. The formation, expansion, and shrinkage of germinal centers are completed approximately 16 days after immunization, inducing the production of antigen-specific terminal plasma cells and memory cells. During somatic hypermutation of central blastema in the dark region of the germinal center, a point mutation occurs in the V domain immunoglobulin gene, resulting in the formation of immunoglobulin with mature affinity 7–10 days after antigen immunization. The selection of antigen-specific central cells occurs in the light zone of the germinal center, a process associated with both recognition and nonrecognition signaling by follicular dendritic cells and T helper cells. Central cells exhibiting antigen-specific, avidity mature sig further differentiate into mature memory cells or terminal plasma cells, while those unselected B cells reenter the dark zone and undergo further somatic mutation and selection processes.

Advantages of RIMMS Technology

There are several immunization strategies that can be used in the preparation of monoclonal antibodies, such as protein immunization, DNA immunization, subtractive immunization, and RIMMS. Compared with other methods, RIMMS has many unique advantages.

The preparation of high-affinity monoclonal antibodies by the RIMMS method requires only 1–2 mice and a small amount of antigen 400ng–3 μg / site or DNA 2.5–100 μg / site.
RIMMS eliminates the need for a blood test during immunization.
Rapid preparation of monoclonal antibodies to various immunogens including synthetic peptides, recombinant proteins, cell extracts, linker complexes, and in vivo DNA expression proteins after immunization has been established.
It is widely used in ELISA, immunoprecipitation, competitive conjugation, immunoblot analysis, immunocytochemistry, or immunohistochemistry studies.

In addition, by combining the method of subtractive immunization, RIMMS, with the methods of DNA immunization and cell immunization, an experimental method for rapid preparation of high-affinity antibodies can be established. This method will greatly shorten the cycle time of monoclonal antibody preparation, which will have a huge impact on the research of proteomics and protein microarrays.

Creative Biolabs is more than willing to discuss with you RIMMS-related techniques and experience to facilitate the smooth running of your research.

All listed services and products are For Research Use Only. Do Not use in any diagnostic or therapeutic applications.

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