title: High-throughput immunoassay for the biochemical diagnosis of Friedreich ataxia in dried blood spots and whole blood.
abstract: BACKGROUND
  Friedreich ataxia (FRDA) is caused by reduced frataxin (FXN) concentrations. A clinical diagnosis is typically confirmed by DNA-based assays for GAA-repeat expansions or mutations in the FXN (frataxin) gene; however, these assays are not applicable to therapeutic monitoring and population screening. To facilitate the diagnosis and monitoring of FRDA patients, we developed an immunoassay for measuring FXN.
  
  
  METHODS
  Antibody pairs were used to capture FXN and an internal control protein, ceruloplasmin (CP), in 15 μL of whole blood (WB) or one 3-mm punch of a dried blood spot (DBS). Samples were assayed on a Luminex LX200 analyzer and validated according to standard criteria.
  
  
  RESULTS
  The mean recovery of FXN from WB and DBS samples was 99%. Intraassay and interassay imprecision (CV) values were 4.9%-13% and 9.8%-16%, respectively. The FXN limit of detection was 0.07 ng/mL, and the reportable range of concentrations was 2-200 ng/mL. Reference adult and pediatric FXN concentrations ranged from 15 to 82 ng/mL (median, 33 ng/mL) for DBS and WB. The FXN concentration range was 12-22 ng/mL (median, 15 ng/mL) for FRDA carriers and 1-26 ng/mL (median 5 ng/mL) for FRDA patients. Measurement of the FXN/CP ratio increased the ability to distinguish between patients, carriers, and the reference population.
  
  
  CONCLUSIONS
  This assay is applicable to the diagnosis and therapeutic monitoring of FRDA. This assay can measure FXN and the control protein CP in both WB and DBS specimens with minimal sample requirements, creating the potential for high-throughput population screening of FRDA.
authors.affiliation: Division of Laboratory Genetics, Department of Laboratory Medicine and Pathology, and.
authors.forename: Devin
authors.initials: D
authors.lastname: Oglesbee
authors.forename: Charles
authors.initials: C
authors.lastname: Kroll
authors.forename: Oleksandr
authors.initials: O
authors.lastname: Gakh
authors.forename: Eric C
authors.initials: EC
authors.lastname: Deutsch
authors.forename: David R
authors.initials: DR
authors.lastname: Lynch
authors.forename: Ralitza
authors.initials: R
authors.lastname: Gavrilova
authors.forename: Silvia
authors.initials: S
authors.lastname: Tortorelli
authors.forename: Kimiyo
authors.initials: K
authors.lastname: Raymond
authors.forename: Dimitar
authors.initials: D
authors.lastname: Gavrilov
authors.forename: Piero
authors.initials: P
authors.lastname: Rinaldo
authors.forename: Dietrich
authors.initials: D
authors.lastname: Matern
authors.forename: Grazia
authors.initials: G
authors.lastname: Isaya
mesh_terms: D000328:Adult; D059788:Dried Blood Spot Testing; D005260:Female; D005621:Friedreich Ataxia / Q000097:blood / Q000175:diagnosis*; D057166:High-Throughput Screening Assays; D006801:Humans; D007118:Immunoassay / Q000379:methods; D007231:Infant, Newborn; D033862:Iron-Binding Proteins / Q000097:blood*; D008297:Male; D012016:Reference Values; D000096923:Frataxin
chemical_list: D033862:Iron-Binding Proteins; D000096923:Frataxin