Possible immune complex interference in the 25OH vitamin D enzyme immunoassay

Abstract

A 42-year-old female with type I diabetes mellitus was found to have renal impairement, gross proteinuria and vitamin D deficiency. Urinary protein excretion rate was 6.29 g/24 hours (<0.22), with a protein/creatinine index of 98.9 (<2.1). Her serum creatinine was 186 µmol/L (60–120), eGFR 28 (>60). Her 25OH vitamin D was undetectable when measured using the IDS enzyme immunoassay (EIA) system. When repeated using LCMS/MS the vitamin D was 41 nmol/L. In addition, rheumatoid factor was negative at <20 (ref range <20) and paraproteins and urinary free light chains were negative. She had positive anti-neutrophil cytoplasmic antibodies (ANCA), anti-proteinase 3 and antinuclear antibody (ANA;titre 1/2560;homogeneous pattern) suggesting systemic lupus disease but negative extractable nuclear antibody (ENA) and dsDNA antibody tests made that diagnosis unlikely. Glomerular basement membrane antibodies, anti myeloperoxidase and cryoglobulins were not detected. Solvent extraction and polyethylene glycol precipitation resulted in vitamin D results of 40 and 68 nmol/L respectively. C3 and C4 were reported at 2.47 g/L (0.6–1.2) 0.86 g/L (0.12–0.35), respectively.Circulating immune complexes may interfere with the EIA. Interferences by immune complexes have been described in immunoassays^1,2 but have not been reported in vitamin D EIAs. As immunoassays are susceptible to interference in serum samples, any unusual results should be verified using another method as a quality assurance practice.