Case Presentation: Functional Assessment of a CASR Variant Identified in a Patient with Hypercalcaemia Confirms Familial Hypocalciuric Hypercalcaemia in the Patient and a Sister Previously Misdiagnosed with Primary Hyperparathyroidism

Figure 1

Western blot analysis of wild type and L34P variant CaSR examined under nonreducing and reducing conditions. HEK293 cells were transfected with wild type or variant CaSR-FLAG-tagged plasmid (5 µg) and then incubated for 48 hrs, and the cells were then lysed, and 80 µg protein, with or without a reducing agent, was separated by SDS-PAGE. The gel was blotted on to nitrocellulose and then probed with mouse monoclonal antibody to FLAG-tag (upper panel) and α-tubulin (as a loading standard; lower panel), followed by goat antimouse HRP-conjugated secondary antibody. Protein bands were visualized using ECL chemiluminescence reagent in a Bio-Rad Molecular Imager. Experimental details are described in reference 18. The image is representative of several independent experiments. The FLAG and tubulin immunodetections were derived from the same blot. Cropping was used to ensure a more concise presentation of the gel image (see Supplementary Figures 1 and 2).