Protocol detail

Immunoblotting Analysis of H+,K+-ATPase Subunits

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H⁺, K⁺-ATPase subunits detection was performed as described previously [15 (link)]. For signaling pathway analysis, PANC-1 cell lysate was boiled 3 min at 95 °C. Primary antibodies against gastric HKα1 (Abcam EPR12251, 1:1000), non-gastric HKα2 (C384-M79, 1:1000, kindly donated by J. J. H. H. M. De Pont and H. G. P. Swarts) [56 (link)], HKβ (Sigma A274, RRID:AB_258027, 1:1000), β-Actin (Santa Cruz Sc-47778, 1:1000, RRID: AB_626632), pSTAT3 (Cell Signaling #9145, 1:1000, RRID: AB_2491009), STAT3 (Cell Signaling #9139, 1:1000, RRID: AB_331757) and Cyclin D1 (Millipore #06-137, 1:500, RRID: AB_2070695) were used.

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Tozzi M., Sørensen C.E., Magni L., Christensen N.M., Bouazzi R., Buch C.M., Stefanini M., Duranti C., Arcangeli A, & Novak I. (2020). Proton Pump Inhibitors Reduce Pancreatic Adenocarcinoma Progression by Selectively Targeting H+, K+-ATPases in Pancreatic Cancer and Stellate Cells. Cancers, 12(3), 640.

Publication 2020

β-Actin Sc-47778

Actin Antibodies Atpase Cell Cyclin d1 Gastric Signaling pathway Stat3 Subunits

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Variable analysis

independent variables

Detection of H+, K+-ATPase subunits

dependent variables

Signaling pathway analysis

control variables

Boiling of PANC-1 cell lysate at 95 °C for 3 minutes

controls

Positive control: None specified
Negative control: None specified

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