Figure 8. ELISA-based uromodulin quantification is not affected by presence of Sda antigen.

(A) Uromodulin serum levels in individuals carrying GG or GA/AA genotype at UMOD variant rs77924615, stratified for their genotype at B4GALNT2 variant rs7224888 (CC, Sda+; TT, Sda-). The expected differences in uromodulin levels are detected regardless of the presence/absence of Sda antigen. The start and end of boxes represent the 25th and 75th percentiles of the uromodulin distribution. The line inside the box represents the median, and the dots indicate outliers above the 75% + 1.5 × interquartile range of uromodulin values. (B) Representative Western blot analysis (left) and relative quantification (right) of uromodulin in lysates of MDCK cells transduced with lentiviral vector expressing HA-tagged uromodulin (lv.HA-hUMOD) and stably expressing B4GALNT2 (Sda+) or not (Sda-). The immunoreactivity of 3 different antibodies (anti-HA, and the 2 antibodies of the Euroimmun ELISA anti-UMOD capture and anti-UMOD detector) was assessed by loading and quantifying increasing amounts of cell lysate. Each value represents the ratio between B11 (negative clone) and B9 (positive clone) expressed as fold relative to the ratio measured with 25 μg of cell lysate (n = 3 independent experiments). Data are represented as vertical scatterplot expressed as mean ± SD (1-way ANOVA; P = 0.66). The ratios obtained for the different antibodies are comparable, suggesting similar immunoreactivity that is not modified by the presence of the Sda antigen.