Utility of Antinuclear Antibody Screening by Various Methods in a Clinical Laboratory Patient Cohort

Abstract

Background Antinuclear antibody (ANA)5 testing is routinely performed during evaluation of patients with a suspected connective tissue disease (CTD), yet the question of which method is most appropriate remains controversial. The purpose of this study was to evaluate the clinical utility of ANA testing by an enzyme immunoassay (EIA), an immunofluorescence assay (IFA), and a multiplex immunoassay (MIA) in a routine laboratory population. Methods Samples (n = 1000) were collected from specimens submitted for ANA testing by EIA (Bio-Rad). All samples were subsequently analyzed by IFA (Zeus) and MIA (Bio-Rad). The sample cohort was weighted to represent the routine testing population. Diagnostic information was obtained by chart review. Results For the diagnosis of a CTD, ROC curve analysis demonstrated no significant differences between IFA (area under the curve 0.81) and EIA (0.84) (P = 0.25), with overlay of a single point for the MIA. When normalized to a specificity of approximately 90%, the sensitivities of the MIA, EIA, and IFA were 67%, 67%, and 56%, respectively. By varying the clinical cutoff, the IFA could achieve the highest sensitivity of 94%; however, the corresponding specificity was only 43%. In contrast, a strongly positive EIA had a specificity of 97%, although, at this cutoff, the sensitivity was only 40%. Conclusions Although the overall diagnostic performance of the IFA, EIA, and MIA were not statistically different, the clinical sensitivity and specificity varied dramatically based on the positive/negative cutoff. Knowledge about the performance characteristics of each method will significantly aid in the interpretation of ANA testing.

In Response to: Utility of Antinuclear Antibody Screening by Various Methods in a Clinical Laboratory Patient Cohort—Differences in Sensitivity, Specificity, and Limitations

“…However, this scenario is rarely the case for the hemolysis/icterus/ lipemia (HIL) 1 index analysis because of its integrated, automated use on the chemistry/immunochemistry platforms. The HIL index analyses seldom has predefined demands to quality performance, meaning that…”

In Response to: Utility of Antinuclear Antibody Screening by Various Methods in a Clinical Laboratory Patient Cohort

“…The authors would like to thank Dr. Baird for his constructive comments regarding the calculation of sensitivity and specificity of antinuclear antibody (ANA) 1 testing in our recent publication (1). The goal of this study was to compare the diagnostic utility of various ANA methods in a representative testing population.…”

Quality Monitoring Approach for Optimizing Antinuclear Antibody Screening Cutoffs and Testing Work Flow

“…This trend is reflected in recent literature and the updated 2015 American College of Rheumatology (ACR) position statement on ANA testing (4,8). Our institution implemented EIA-based ANA screens in July of 2015 as a way to focus manual IFA testing on specimens more likely to be positive.…”

“…IFA using HEp-2 substrate is considered the gold standard method for clinical ANA testing (2)(3)(4); it is historically a manual technique with some automation recently available in the US market. MIA and EIA are generally more automatable than IFA and have variable sensitivity for CTD depending on how they are configured (2,3,(5)(6)(7)(8).…”