Stability of serum precipitins to Aspergillus fumigatus for the diagnosis of allergic bronchopulmonary aspergillosis

Abstract

Background: Allergic bronchopulmonary aspergillosis (ABPA) reflects hypersensitivity and an exaggerated immune response to Aspergillus fumigatus. ABPA typically occurs in individuals with airway diseases such as asthma or cystic fibrosis and is associated with worse outcomes for individuals with these conditions. Each year, physicians across the province of British Columbia submit over 2600 diagnostic testing requests to a centralized location in Vancouver, requiring specimen collection, storage, and shipment from different clinics across the province. Timely and reliable testing of Aspergillus precipitins is critical to optimizing diagnosis and management of ABPA. At our centre, we analyzed sample stability in varying storage conditions to provide guidance to those using this routine diagnostic test. Methods: To determine temperature and time stability, 31 serum specimens positive for Aspergillus fumigatus precipitins from routine clinical testing were each aliquoted and incubated at 4 and 37 °C. Samples were repeatedly assayed for precipitins to Aspergillus fumigatus via agarose gel double immunodiffusion (AGID) at 7, 14, and 28 days post-incubation. To determine freeze-thaw stability, 39 serum specimens submitted for routine clinical testing for Aspergillus precipitins were randomly selected. Each specimen was aliquoted and stored at 4 or -20 °C. 4 °C samples were maintained at 4 °C while -20 °C samples were split into three groups corresponding to one, two, or three freeze-thaw cycles. -20 °C samples were thawed at room temperature in the morning and then immediately frozen overnight for up to a total of three freeze-thaw cycles. Results: Regarding temperature and time stability, median stability time was 47 and 34 days at 4 and 37 °C, respectively. The log-rank model indicates no statistically significant difference between the two temperature storage conditions (p = 0.14) with a hazard ratio of 0.61 (95% CI, 0.31-1.2). In terms of freeze-thaw stability, no indication of serum degradation with regards to Aspergillus fumigatus precipitins was found with repeated freeze-thaw cycles as compared to refrigerated storage. Conclusions: The stability of serum precipitins to Aspergillus fumigatus was found to be dependent on time, but not temperature and freeze-thaw cycles. Specimens for Aspergillus fumigatus precipitins testing should be shipped at ambient temperature and tested within 2 weeks from collection.