The Immunsystem‑to‑go project, funded by the German Federal Ministry of Education and Research (grant 031B0818A) and carried out from 1 July 2019 to 30 April 2023, aimed to create a next‑generation, antibody‑free platform for rapid detection of microorganisms in drinking water. The core concept was to use human innate immune receptors, Toll‑like receptors (TLRs), which bind conserved microbial components such as lipopolysaccharide (LPS) and lipoproteins. By exploiting the high affinity of TLRs for these ligands, the system could recognise entire classes of bacteria—gram‑positive or gram‑negative—without the need for specific antibodies or lengthy culture steps.
A key technical milestone was the development of two membrane‑based assay formats: a lateral‑flow assay (TLR‑LFA) and a flow‑through assay (TLR‑FTA). Initial attempts to produce mobile TLRs from recombinant HEK cells supplied by the Fraunhofer Institute for Interface and Bioprocess Engineering (IGB) were hampered by supply constraints. Consequently, the consortium turned to the antimicrobial peptide polymyxin B (PMB), which binds LPS with very high affinity. PMB was incorporated as a capture ligand in both the TLR‑LFA and TLR‑FTA formats. Concentration‑dependent binding of PMB to E. coli, purified LPS, and Legionella pneumophila was demonstrated in ELISA and lateral‑flow formats, confirming that the ligand could effectively replace TLRs for Gram‑negative detection.
The project validated the PMB‑based TLR‑FTA against conventional, time‑consuming Legionella culture methods. The prototype assay achieved sensitivities and specificities comparable to the established tests, indicating that the new platform can deliver rapid, reliable results. In parallel, a fluorescence‑based detection device was prototyped, combining a fluorophore‑labeled probe with a custom detector. This system was integrated into a 3D‑printed housing, allowing for modular assembly and potential adaptation to other assay formats.
Additional technical achievements included the creation of an activity assay for the TLR4/MD2 complex, enabling functional verification of receptor preparations. The consortium also developed a range of assay components that can be repurposed for other microbial targets, demonstrating the platform’s versatility. The 3D‑printed housings and components for the FTA and analysis device were produced in a 0‑series, laying the groundwork for future production runs.
Collaboration was structured around four partners: NH DyeAGNOSTICS GmbH (NHD) led the integration of PMB into the assay work packages and oversaw the development of the fluorescence detector; the Fraunhofer Institute (IGB) was responsible for providing functional TLRs and initial recombinant expression; Senova Gesellschaft für Biowissenschaften und Technik GmbH (SEN) contributed to ligand screening and assay optimisation; and fzmb GmbH (Forschungszentrum für Medizintechnik und Biotechnologie) supplied expertise in biotechnological process development. NHD handled work packages AP 0, 1, 3, 5, 9, 10, 11, and 12, coordinating the overall project timeline and deliverables.
In summary, the Immunsystem‑to‑go project successfully established a TLR‑based detection platform that bypasses the need for antibodies and culture, achieving performance metrics on par with conventional Legionella assays. The use of PMB as a surrogate ligand, coupled with fluorescence readout and 3D‑printed hardware, provides a scalable, rapid testing solution that can be extended to other microbial groups. The collaborative effort among German research institutes and industry partners, supported by federal funding, has produced a validated prototype and a clear pathway toward commercial deployment.