Development and clinical application of a highly sensitive serological assay for detecting thrombospondin type-1 domain-containing 7A antibodies in idiopathic membranous nephropathy.

OBJECTIVE

To establish a highly sensitive serological assay for detecting serum thrombospondin type-1 domain-containing 7 A antibodies (THSD7A-Ab) in patients with idiopathic membranous nephropathy (IMN), thereby addressing the current gap in available diagnostic biomarkers.

METHODS

A time-resolved fluorescence immunoassay (TRFIA) was established by immobilizing THSD7A protein on the solid phase, which captured THSD7A-Ab present in serum samples. Detection was performed using a europium (Eu3+)-labeled secondary antibody. Serum THSD7A-Ab levels were measured in both THSD7A-positive IMN patients and M-type phospholipase A2 receptor (PLA2R)-negative IMN patients, and the corresponding seropositivity rates were determined.

RESULTS

The TRFIA-THSD7A-Ab assay demonstrated a sensitivity of 0.13 RU/mL, with a linear range from 0.13 to 600 RU/mL. The intra-assay precision ranged from 3.35% to 4.63%, while the inter-assay precision ranged from 5.21% to 9.07%. The recovery rate was 93.78% to 107.20%. Serum THSD7A-Ab levels in THSD7A-associated IMN patients (176.54 ± 224.77 RU/mL) were significantly higher PLA2R-associated IMN patients (11.38 ± 4.76 RU/mL, P < 0.0001) and healthy controls (7.86 ± 2.92 RU/Ml, P < 0.0001). The cut-off value for distinguishing THSD7A-associated IMN was determined to be 13.70 RU/mL. Among PLA2R-negative IMN patients, the positive rate of THSD7A-Ab was 1.6%.

CONCLUSION

A highly sensitive TRFIA method for detecting serum THSD7A-Ab was successfully established for the first time, demonstrating excellent discriminative capability for identifying THSD7 A-positive patients.