کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1243338 | 1495797 | 2015 | 6 صفحه PDF | دانلود رایگان |
• We used an strong cation exchange monolith for large volume sample preconcentration in CZE-based proteomics.
• We used a pH junction to concentrate peptides after elution from the monolith.
• We observed a 3000-fold enrichment for angiotensis II.
• We identified over 100 proteins from a 1 µg/mL tryptic digest of E. coli.
A sulfonate–silica hybrid strong cation-exchange (SCX) monolith was synthesized at the proximal end of a capillary zone electrophoresis column and used for on-line solid-phase extraction (SPE) sample preconcentration. Sample was prepared in an acidic buffer and deposited onto the SCX-SPE monolith and eluted using a basic buffer. Electrophoresis was performed in an acidic buffer. This combination of buffers results in formation of a dynamic pH junction, which allows use of relatively large elution buffer volume while maintaining peak efficiency and resolution. All experiments were performed with a 50 µm ID capillary, a 1 cm long SCX-SPE monolith, a 60 cm long separation capillary, and a electrokinetically pumped nanospray interface. The volume of the capillary is 1.1 µL. By loading 21 µL of a 1×10−7 M angiotensin II solution, an enrichment factor of 3000 compared to standard electrokinetic injection was achieved on this platform while retaining efficient electrophoretic performance (N=44,000 plates). The loading capacity of the sulfonate SCX hybrid monolith was determined to be ~15 pmol by frontal analysis with 10−5 M angiotensin II. The system was also applied to the analysis of a 10−4 mg/mL bovine serum albumin tryptic digest; the protein coverage was 12% and 11 peptides were identified. Finally, by loading 5.5 µL of a 10−3 mg/mL E. coli digest, 109 proteins and 271 peptides were identified in a 20 min separation; the median separation efficiency generated by these peptides was 25,000 theoretical plates.
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Journal: Talanta - Volume 138, 1 June 2015, Pages 117–122