Cincinnati Children's Hospital Medical Center Cincinnati, Ohio, United States
No financial relationships with ineligible companies to disclose
Ekemini Ogbu1, Stela Florea2, Donna Diorio2, Somak Roy2, Dhriti Sharma2, Michael Henrickson2, Patricia Vega-Fernandez2, Angela Migowa3, Sarangarajan Ranganathan2 and hermine brunner1, 1Cincinnati Children's Hospital Medical Center, University of Cincinnati College of Medicine, Cincinnati, OH, 2Cincinnati Children's Hospital Medical Center, Cincinnati, OH, 3AGA KHAN UNIVERSITY, Nairobi, Kenya Background/Purpose: Timely diagnosis of children and young adults with rheumatologic disorders remains a global challenge especially in lower-resource settings.There is limited access to diagnostic laboratory tools such as antinuclear antibody (ANA) testing which is necessary to confirm a diagnosis of several rheumatic disorders. Easier availability of ANA testing will allow for timely initiation of life-saving treatment and improve outcomes. Thus, our objective was to develop a lower cost, accurate and scalable dried blood spot (DBS) ANA testing option that can be effectively used in lower-resource settings in Africa and other parts of the world. Methods: De-identified cross-sectional blood samples collected via venipuncture were tested for the presence of ANA by the gold standard immunofluorescence (IIF) and ELISA on DBS (ELISA-DBS). For ELISA-DBS, serum or whole blood (WB) samples were transferred to a protein-optimized DBS paper matrix and eluted overnight prior to ELISA testing (measured as positive or negative). Initially, the presence of ANA in serum samples was tested concurrently by IIF (1:80 – 1:1280 titers) and ELISA-DBS. Following this, paired serum and WB samples of varying ANA titers by IIF (1:80 – 1:2560) were also tested by ELISA-DBS. Then, ANA results by IIF and by ELISA-DBS tests were compared for varying (10 – 75 microliter) and stable (30 microliter) DBS sample quantities for volume and stability studies respectively. Comparison of ELISA-DBS from paired serum and WB samples was also performed following exposure of DBS samples to room temperature and 370C, and on prolonged DBS sample storage (24hours/7days/14days) prior to ELISA-DBS testing. Results: Comparison of serum samples concurrently tested for ANA by IIF and by ELISA-DBS showed 80% concordance (N = 4, Table 1). There was 100% concordance of paired serum and WB samples of varying ANA titers by IIF with ELISA-DBS (N= 3, Table 2). ELISA-DBS from paired 30 microliter serum and WB samples at 24hrs/7days/14days yielded 83/100/67% concordance in ANA results respectively (Figure 1). Conclusion: Our pilot study demonstrates that ANA testing from DBS is feasible with good concordance at varying ANA titers and with temperature excursions. Our novel ANA testing by ELISA-DBS could aid earlier diagnoses of rheumatic diseases in lower resource settings.
Table 1: Comparison of ANA Testing by Immunofluorescence and by ELISA on Dried Blood Spot from Serum Samples
Table 2: Comparison of ANA Testing by Conventional ELISA and ELISA on Dried Blood Spot from Paired Serum and Whole Blood Samples
Figure 1: Positive Concordance of ELISA-DBS from Serum and Whole Blood Re-measured on Day 7
E. Ogbu: None; S. Florea: None; D. Diorio: None; S. Roy: None; D. Sharma: None; M. Henrickson: None; P. Vega-Fernandez: None; A. Migowa: None; S. Ranganathan: None; h. brunner: AbbVie/Abbott, 2, AstraZeneca-Medimmune, 2, Biogen, 2, Boehringer-Ingelheim, 2, Bristol-Myers Squibb, 2, 12, Contributions, Celgene, 2, Eli Lilly, 2, 12, Contributions, EMD Serono, 2, F. Hoffmann-La Roche, 2, 12, Contributions, GlaxoSmithKlein(GSK), 2, 6, 12, Contributions, Janssen, 2, 12, Contributions, Merck/MSD, 2, Novartis, 2, 6, 12, Contributions, Pfizer, 2, 12, Contributions, Roche, 6, R-Pharm, 2, Sanofi, 2, UCB, 2.