A Novel Monoclonal Antibody Detects Aspergillus Galactomannan Antigens in Urine

K. Datta, S. F. Dufresne, X. Li, M. Feldmesser, J. F. Staab, K. A. Marr


Invasive mold infections are a major cause of morbidity and mortality in immunocompromised patients, including bone-marrow transplant recipients and those with acute hematological malignancies. Culture and histopathology are currently the gold standards of diagnosis, but are time-consuming, lack sensitivity, and often require samples to be retrieved by invasive and hazardous procedures. This accounts for the increasing usage of culture-independent diagnostic methods. The single established commercial immunoassay detecting the Aspergillus antigen, galactomannan (GM), is currently FDA-cleared for serum and broncho-alveolar lavage specimens only. However, urine would be a desirable alternative; it is amenable to point of care (POC) testing, and known to contain GM-like polysaccharide antigens in the course of several invasive fungal infections, including invasive aspergillosis (IA). A sandwich ELISA using a novel monoclonal antibody (MAb 476, a mouse IgMκ generated against A. fumigatus) was able to detect polysaccharide exoantigens from A. fumigatus strain AF293, both an ethanol-precipitable (EP) fraction of culture supernatant, and chemically purified GM. In addition, the assay recognized EP antigens from a panel of clinically relevant molds, including various species of Aspergillus (except A. terreus) and Fusarium. This ELISA was able to detect Aspergillus antigen in simulated human urine samples containing either GM or EP antigen. We observed that urine interfered with the immunological reaction, which could not be attributed to a particular component or physicochemical parameter. However, this interference could be alleviated by removing small molecular weight substances. We could also increase the sensitivity of the assay by concentrating the sample prior to desalting, and were able to detect Aspergillus antigen in several urine samples from patients with proven IA. In this study, we have characterized (a) a novel MAb capable of recognizing GM-like polysaccharide exoantigens from different Aspergillus species, and (b) an ELISA to detect Aspergillus GM antigens in simulated and patient urine samples. Efforts are currently underway to port this antibody to a different platform suitable for a POC device.

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American Society for Microbiology General Meeting
    • ASM 112th (2012)