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ABSTRACT: A method for integrating nanoelectrospray mass spectrometry with a microreactor for on-line digestion and fast peptide mass mapping from dilute protein samples is presented. Fused silica capillaries (i.d. 50 microm, o.d. 360 microm) are employed as the digestion microreactor and the nanoelectrospray emitter by immobilizing trypsin onto the surface of the inner wall of the fused silica capillary tubing. The procedure is demonstrated using solutions of 1pmol/mul angiotensin II, cytochrome c, hemoglobin, and beta-casein. Because the inner walls of the capillaries are modified by covalent chemical bonds, the adsorption of peptides and proteins to the inner walls of the capillaries is suppressed. This procedure was performed with solutions as dilute as 1fmol/mul (1nM) cytochrome c. This method shows generation of tryptic peptides with sequence coverage up to 90% within minutes; trypsin autolysis products are not detected. In addition, the immobilized enzyme can be cleaned easily, enabling the microreactor to be reused for nanoelectrospray.
Analytical Biochemistry 01/2007; 359(2):167-75. · 3.00 Impact Factor
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ABSTRACT: As miniaturization of electrospray continues to become more prevalent in the mass spectrometry arsenal, numerous types of conductive coatings have been developed with miniaturized electrospray emitters. Different conductive coatings have different properties that may lead to differences in analytical performance. This paper investigates and compares the analytical properties of a series of applied conductive coatings for low-flow electrospray ionization developed in this laboratory vs. commercially-available types. Evaporated graphite is thoroughly compared with commercially available polyaniline (PANI) coated emitters and metal coated emitters. Each set of emitters was investigated to determine various performance characteristics, including susceptibility to electrical discharge in both positive and negative ionization modes, as well as emitter reproducibility and generation of a standard curve to determine each emitter coating's limit of detection and limit of quantitation. Furthermore, evaporated graphite and polyaniline coated fused silica capillaries were investigated to determine which coating is more stable over long-term analyses and during electrical discharge.
The Analyst 05/2006; 131(4):547-55. · 4.23 Impact Factor
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ABSTRACT: A new, more rapid method for coating nanoelectrospray emitters with graphite is to use a vacuum deposition chamber and a graphite carbon electrode. This method allows for mass production of nanoelectrospray emitters in a short period of time. The emitters are laser-pulled borosilicate glass micropipets and have tapers of around 4 microm i.d. The conductive coating applied to the emitter is only 20-30 nm thick, allowing for optical transparency with the borosilicate emitters. The conductive coating is stable for a number of hours at the high voltages used for nanoelectrospray ionization and is durable in both positive and negative ion modes-even during electrical discharge. This stability will make it possible to couple these emitters with online separations such as capillary liquid chromatography or capillary electrophoresis.
Analytical Chemistry 01/2004; 75(24):7015-9. · 5.86 Impact Factor
