A “plug-and-use” approach towards facile fabrication of capillary columns for high performance nanoflow liquid chromatography

• NanoLC columns can be fabricated in a facile “plug-and-use” approach.
• Superb performance consistency at a column-to-column level was demonstrated.
• Consistently high peak capacity for protein digest separations was achieved.

Capillary columns used for nanoflow liquid chromatography play an important role in modern proteomics. High quality columns are needed to provide high peak capacity and highly reproducible separations. This is extremely important when multiple separations were compared in parallel in searching for potential biomarkers. Herein, we introduce a “plug-and-use” fritting technology for fabrication of high quality and highly reproducible capillary columns. Due to the identical length, good permeability, and stability of the prefabricated frits adopted, the capillary columns presented excellent performance consistency in terms of retention time, peak width as well as peak capacity at a column-to-column level (relative standard deviations, RSDs, at 0.4–0.9%, 2.1–3.6%, and 2.7%, respectively, n = 6) for separations of complex mixtures of protein digest. For capillary columns packed with 5 μm particles, high separation efficiency was demonstrated by the minimum plate height of 11 μm, approaching the theoretical performance limit of such material. For separations of protein digests, the columns demonstrated excellent peak capacities of 110 and 300 for 20 and 360 min gradients, respectively. The simple fabrication, good performance as well as consistent quality of such columns provide a reliable tool for high throughput separations requiring the use of multiple high performance capillary columns in parallel.