LegioTyper: Rapid typing of Legionella pneumophila serogroup 1 by flow-based chemiluminescence sandwich microarray immunoassay

Chemiluminescence imaging of a sandwich microarray immunoassay (CL-SMIA) is a simple and rapid method for antibody-based serotyping of pathogens like L. pneumophila, which is important to identify even outbreak-associated subtypes like L. pneumophila Sg 1 subtype Philadelphia. The CL image pattern can be interpreted by a decision path similar to the well-known "Dresden Panel", which is typically performed via indirect ELISA. New antibodies needed to be characterized for the so-called LegioTyper panel as capturing antibodies to perform the flow-based CL-SMIA. The following was achieved for the LegioTyper principle: (i) minimizing the costs for antibody microarrays by manufacturing Jeffamine-2003-coated polycarbonate sheets, (ii) optimized automated test procedure which is able to perform the CL-SMIA in 34 min, and (iii) covalent immobilization of cAb using a microdispensing buffer which contains reactions for stabilizing and bioconjugation of cAb. Nine mAb and one polyclonal Ab (pAb) were characterized regarding the selectivity to L. pneumophila Sg 1 and additionally to the virulence-associated subtypes Knoxville, France, Allentown, Benidorm, and Philadelphia (all Pontiac group), OLDA, Oxford, Bellingham, Heysham, and Camperdown (all non-Pontiac group). We could show that by using the LegioTyper it is possible to differentiate between L. pneumophila Sg1 and non-Sg1 and between Pontiac and non-Pontiac strains. Additionally, subtyping with the "LegioTyper Panel" was possible for most of the outbreak-associated subtypes. Overall, our study showed that the LegioTyper principle can be used for rapid antibody-based subtyping of L. pneumophila Sg 1 after cultivation. It is an alternative to indirect ELISA or qPCR, which are more labor- and cost-intensive, and the automation is more complicated.