Monoclonal antibodies to hyphal exoantigens derived from the opportunistic pathogen, Aspergillus terreus.
Nayak-AP; Green-BJ; Janotka-E; Hettick-JM; Friend-S; Vesper-SJ; Schmechel-D; Beezhold-DH
Clin Vaccin Immunol 2011 Sep; 18(9):1568-1576
A. terreus has been difficult to identify in cases of aspergillosis and clinical identification has been restricted to the broad identification of aspergillosis lesions in affected organs or the detection of fungal carbohydrates. As a result, there is a clinical need to identify species-specific biomarkers that can be used to detect invasive A. terreus disease. Monoclonal antibodies (mAbs) were developed to a partially purified preparation of cytolytic hyphal exoantigens (HEA) derived from A. terreus culture supernatant (CSN). Twenty-three IgG1 isotype murine mAbs were developed and tested for cross reactivity against hyphal extracts of 54 fungal species. Sixteen mAbs were shown to be specific for A. terreus. HEA antigens were detected in conidia, hyphae and in CSN of A. terreus. HEA antigens were expressed in high levels in the hyphae during early stages of A. terreus growth at 37°C, whereas at room temperature, the expression of HEA antigens peaked by day 4-5. Expression kinetics of HEA antigens in CSN showed a lag, with peak levels at later time points at RT and 37°C compared to hyphal extracts. Serum spiking experiments demonstrated that human serum components do not inhibit detection of the HEA antigen epitopes by mAb ELISA. Immunoprecipitation and proteomic analysis demonstrated that mAbs 13E11 and 12C4 immunoprecipitated a putative uncharacterized leucine aminopeptidase (Q0CAZ7), while mAb 19B2 recognized a putative dipeptidyl-peptidase V (DPP5). Studies using Confocal laser scanning microscopy showed that the uncharacterized leucine aminopeptidase mostly localized to extracellular matrix structures, while dipeptidyl-peptidase V was mostly confined to the cytoplasm.
Antibody-response; Antigens; Biomarkers; Cell-biology; Cytology; Fungal-infections; Fungi; Growth-factors; Growth-rate; Humans; Immune-reaction; Immunology; Laboratory-techniques; Laboratory-testing; Microbiology; Statistical-analysis; Temperature-effects; Temperature-measurement;
Author Keywords: Aspergillus terreus; immunoglobulin G; monoclonal antibody; exoantigen
Donald H. Beezhold, Allergy and Clinical Immunology Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention, Morgantown, WV
Healthcare and Social Assistance; Services
Clinical and Vaccine Immunology