Journal: Scientific Reports

Article Title: The two-pore channel TPC1 is required for efficient protein processing through early and recycling endosomes

doi: 10.1038/s41598-017-10607-4

Figure Lengend Snippet: Immunohistochemical detection of TPC1 in kidney and co-localization with syntaxin-12. ( a ) TPC1 (red) is predominantly expressed in the proximal tubule as indicated by co-staining with the marker Lotus tetragonolobus lectin (green) (Vector laboratories #B1325). ( b ) Control staining using a kidney section from a TPC1 knock-out mouse. Staining with antibodies as in ( a ), scale bar = 100 µm for ( a ) and ( b ). ( c ) Co-localization of TPC1 (red, anti-TPC1, generated by Gramsch Laboratories) with syntaxin-12 (green, anti-STX12 Synaptic Systems) at the apical membrane of kidney proximal tubules. DAPI staining in blue; scale bar = 20 µm.

Article Snippet: For antigen retrieval kidney cryosections were treated with citrate buffer (pH 6, microwave heating for 10 min) and then incubated with blocking solution (phosphate buffer containing 1% Triton X-100, 1% bovine serum albumin and 5% normal goat serum) for 1–2 h. Sections were incubated for 3 h at room temperature with either the anti-TPC1 antibody (5 µg/ml in blocking solution) and anti-Syntaxin-12 (monoclonal mouse, Synaptic Systems GmbH; 2 µg/ml in blocking solution) or anti-TPC1 and biotinylated Lotus tetragonolobus lectin (LTL, 10 µg/ml, FITC-conjugated from Vector Laboratories) followed by a 4 °C overnight incubation.

Techniques: Immunohistochemical staining, Staining, Marker, Plasmid Preparation, Knock-Out, Generated