Laboratory testing for anti-synthetase antibodies: a novel algorithm to improve diagnostic accuracy

Area of Science:

• Clinical immunology
• Autoimmune diagnostics
• Laboratory medicine

Background:

• Anti-synthetase syndrome (ASyS) is characterized by autoantibodies against aminoacyl-tRNA synthetases (ARS).
• Existing commercial assays for anti-ARS antibodies exhibit variable accuracy.
• This study addresses the need to evaluate commonly used anti-ARS assays in Spanish clinical labs.

Purpose of the Study:

• To evaluate the accuracy and reliability of commercial anti-ARS assays.
• To identify methods for improving the diagnostic performance of anti-ARS detection.
• To develop a diagnostic algorithm for more dependable anti-ARS testing.

Main Methods:

• Evaluated 99 serum samples from five Spanish laboratories, initially positive for anti-ARS by line blot (LB).
• Samples were re-tested using indirect immunofluorescence (IIF) on HEp-2 cells, dot blot (DB), and RNA immunoprecipitation (RNA-IP) at a central laboratory.
• Confirmation of anti-ARS antibodies was based on DB or RNA-IP results.

Main Results:

• Only 27% of initial anti-ARS positive samples were confirmed, with a 5% confirmation rate for LB intensity 1+ and 2+.
• Increasing the LB cut-off to 3+ and including a diagnosis-compatible IIF pattern achieved a 100% confirmation rate.
• RNA-IP was most sensitive for anti-PL-12, anti-EJ, and anti-OJ; DB was more sensitive for anti-Jo-1 and anti-PL-7.

Conclusions:

• Commercial LB methods for anti-ARS detection show significant variability, with a high unconfirmed rate (73%).
• A LB signal intensity cut-off of 3+ combined with a compatible IIF pattern substantially increases specificity.
• A new diagnostic algorithm can enhance the reliability of anti-ARS detection in routine practice.