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Aim:We compared four commonly used, commercially available reverse phase nanoLC columns for identification/determination of Wnt/β-catenin-related pathway proteins.Materials & methods:The columns were: Chromolith® (silica monolith; Merke Millipore, MA, USA), PepMap™ (porous particles; Thermo Fisher Scientific, MA, USA), Accucore™ (solid core particles; Thermo Fisher Scientific) and PepSwift™ (organic monolith; Thermo Fisher Scientific).Results:The peak capacity of the columns varied from 100 (Pepswift) to 190 (Accucore) (for 30 min gradients). All columns enabled identification/detection of GSK3β and β-catenin in the complex samples. However, even the columns with higher peak capacities could not enable detection of the somewhat less abundant proteins AXIN2 and TNKS2. The monoliths were more prone to retention time instability when sample complexity increased.Conclusion:We find that commercial nanoLC columns, although featuring different morphologies and peak capacities, provided surprisingly few practical differences for relatively fast, targeted determination of proteins.