Novel Monoclonal Antibodies 1D2 and 4E4 Against <i>Aspergillus</i> Glycoprotein Antigens Detect Early Invasive Aspergillosis in Mice

Due to the high morbidity and mortality rates of invasive aspergillosis (IA) and the importance of early IA detection for successful treatment and subsequent outcome, this study aimed to determine a time course of detectable antigen in a mouse model of IA and correlate it with tissue invasion by using two novel monoclonal antibodies, 1D2 and 4E4, that can be used to detect the <i>Aspergillus</i>-derived glycoproteins. Immunocompromised mice were randomly divided into five groups: uninfected control, and inoculation with conidia from <i>Aspergillus fumigatus, Aspergillus flavus</i>, <i>Aspergillus niger</i>, and <i>Aspergillus terreus</i>. Conidia (2 × 10⁶ cells/mL) were administered intravenously via tail vein injection. Three mice from each group were euthanised at each time point (6 h, 12 h, 18 h, 24 h, and 48 h) after inoculation. Urine and blood were collected for analysis using a double-sandwich ELISA using 1D2 and 4E4. Liver, spleen, and kidney tissues were harvested for tissue staining. The levels of liver injury in the IA mice progressively increased with time after inoculation with <i>Aspergillus</i> conidia. Following inoculation with <i>A. fumigatus</i>, swollen conidia were identified in the spleen, as well as antigens in blood after 18 h. Hyphae were detected in the spleen, liver, and kidney after 48 h. For <i>A. flavus,</i> the antibodies detected hyphae in the liver and spleen as well as circulating antigens in blood samples 48 h after inoculation. Tissue injury was observed in the mice inoculated with <i>A. terreus</i> and <i>A</i>. <i>niger</i>, but there was no evidence of fungal invasion or antigens in the blood. Antigens were not detectable in mouse urine but could be detected in glomeruli of the kidney by immunofluorescence. In conclusion, the mAb-based antigen detection double-sandwich ELISA results were consistent with the IHC results in this study. Novel monoclonal antibodies 1D2 and 4E4 can serve as tools for the early identification of IA in mice infected by <i>A. fumigatus</i> and <i>A. flavus</i>. This study also suggests the potential usefulness of this approach in human disease.