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RAPPID-M: A Mix-and-Measure Bioluminescent Sandwich Immunoassay Based on Generic Antibody-Binding Protein M
Anna Swietlikowska1,2, Eva van Aalen1,2, Max Bossink1,2
1Laboratory of Chemical Biology, Department of Biomedical Engineering, Eindhoven University of Technology, PO Box 513, Eindhoven 5600 MB, The Netherlands.
ACS Sensors
|October 10, 2025
Summary
A new RAPPID-M sensor platform enables homogeneous immunoassays using protein M, expanding compatibility to more antibody types like mouse IgG1 and antibody fragments. This innovation simplifies the assay process and enhances analytical performance for biomarker detection.
Area of Science:
- Biotechnology
- Assay Development
- Immunochemistry
Background:
- Homogeneous immunoassays offer advantages over heterogeneous assays by eliminating washing steps.
- The original RAPPID platform utilized split luciferase complementation for solution-based immunoassays but required protein G-mediated photoconjugation.
- Protein G conjugation limits compatibility with certain antibody isotypes, such as mouse IgG1.
Purpose of the Study:
- To develop a next-generation RAPPID sensor platform (RAPPID-M) with broader antibody compatibility.
- To eliminate the need for protein G and photo-cross-linking in RAPPID assays.
- To improve the ease of use and analytical performance of homogeneous immunoassays.
Main Methods:
- Developed RAPPID-M sensors employing a generic antibody-binding domain from protein M.
- Utilized protein M for irreversible binding to antibody fragments and various IgG isotypes.
- Investigated the compatibility of RAPPID-M with mouse IgG1, scFv, and Fab antibody fragments.
Main Results:
- RAPPID-M demonstrates compatibility with protein G-incompatible mouse IgG1 antibodies, as well as scFv and Fab antibody fragments.
- The irreversible binding of protein M eliminates the requirement for photo-cross-linking.
- RAPPID-M offers a mix-and-measure assembly, simplifying conjugate preparation.
- Analytical performance was maintained and in some cases improved compared to the original RAPPID platform.
Conclusions:
- RAPPID-M represents a significant advancement in homogeneous immunoassay technology.
- The platform's enhanced compatibility and simplified workflow broaden its applicability in biomarker detection.
- RAPPID-M provides a robust and versatile tool for sensitive and efficient immunoassays.
