Laboratory assessment
Antinuclear antibodies detection by the indirect immunofluorescence
assay (IIFA) was performed using HEp-2 cells (Menarini, Florence,
Italy) and antigen specificity was determined by ELISA test.
Erythrocyte sedimentation rate (ESR), CRP, C3, uric acid, serum
creatinine (sCr) and urinalysis were evaluated.
The serum and urine were collected for the determinations of FLC
concentrations on OPTILITE instruments (The Binding Site Group Ltd,
Birmingham, UK). The quantitative detection of FLC has been obtained
using the latex particle enhanced Freelite® k and λ immunoassays (The
Binding Site Group Ltd, Birmingham, UK) following the manufacturer’s
instructions.
The lower limit of quantification was 0.1 mg/L. The immunoassay
consisted of two separate measurements in serum, for free k (normal
range: 3.3-19.4 mg/L) and free λ (normal range: 5.7-26.3 mg/L). A ratio
of k/λ <0.26 or >1.65 is abnormal, according to
the manufacturer’s recommendations. In urine, the normal value of free k
is 0.39–15.1 mg/L and free λ is 0.81–10.1 mg/L. A ratio of k/λ uFLC
>1.80 is considered abnormal. Samples were thawed only once
and immediately assayed in a single batch, following the manufacturer’s
instructions. All the determinations were performed by an operator
without knowledge of the clinical information of the handled sample.
Each sample was tested twice to minimize eventual discrepancies, and all
tests were performed in the same laboratory with the same instruments.