Micro-ESI

Utilization of capillary columns in conjunction with micro ESI devices is becoming a new trend in the field of LC/MS. Capillary HPLC has become a particularly important technique in situations where the supply of analyte is limited, such as in proteomic analysis. According to studies conducted by Smith et al., only one in a hundred thousand of analyte molecules present in solution eventually reach mass detection in a conventional ESI interface. Smith et al. attributed this poor electrospray... 

Martin, S. E., Shabanowitz, J., Hunt, D. R, and Marto, J. A., Subfemtomole MS and MS/MS peptide sequence analysis using nano-HPLC micro-ESI Eourier transform ion cyclotron resonance mass spectrometry, Analytical Chemistry 72(18), 4266 274, 2000. 

The ability of this ionization method for the determination of very high molecular weights is illustrated in Figure 1.26 [68], The spectrum displayed is obtained from assemblies of vanillyl alcohol oxidase containing respectively 16 and 24 proteins. The spectrum was obtained with a hybrid quadrupole TOF instrument, Q-TOF Micromass, equiped with a micro-ESI source. To obtain such a spectrum one needs not only a mass spectrometer with sufficient mass range and resolution, but also high skill in protein purification. 

M.R. Emmett, R.M. Caprioli, Micro-ESI-MS ultra-high-sensitivity analysis of peptides and proteins, J. Am. Soc. Mass Spectrom., 5 (1994) 605. 

D. Figeys, Y. Ning, R. Aebersold, A microfabricated device for rapid protein identification by micro-ESI—ion-trap MS, Anal. Chem., 69 (1997) 3153. 

From a practical point of view, the discussion on flow-rate can be summarized as follows. In LC-APCI-MS, the typical flow-rate is 0.5-1.0 ml/min. For routine applications of LC-ESI-MS in many fields, extreme column miniaturization comes with great difficulties in sample handling and instrument operation. In these applications, LC-MS is best performed with a 2-mm-ID column, providing an optimum flow-rate of 200 pFmin, or alternatively with conventional 3-4.6-mm-ID columns in combination with a moderate split. In sample limited cases, further reduction of the column inner diameter must be considered. Packed microcapillary and nano-LC columns with micro-ESI and nano-ESI are rontinely applied inproteomics stndies (Ch. 17.5.2). 

E. Hoyes, S.J. Gaskell, Tutowahc function switching and its usefulness in peptide and protein analysis using direct infusion micro-ESI Q-TOF MS, Rapid Commun. Mass Spectrom., 15 (2001) 1802. 

C.L. Gatlin, G.R. Kleemann, L.G. Hays, A.J. Link, J.R. Yates, HI, Protein identification at the low femtomole level from silver-stained gels using a new fritless ESI interface for LC-micro-ESI- and nano-ESI-MS, Anal. Biochem., 263 (1998) 93. 

C.L. Gatlin, J.K. Eng, S.T. Cross, J.C. Detter, J.R. Yates, III, Automated identif cation of amino acid sequence variations in proteins by HPLC-micro-ESI-MS-MS, Anal. Chem., 72 (2000) 757. 

F. Zappacosta, M.J. Huddleston, R.L. Karcher, V.I. Gelfand, S.A. Carr, R.S. Annan, Improved sensitivity for phosphopeptide mapping using capillary LC and micro-ESI-MS comparative phosphorylation site mapping from gel-derived proteins. Anal. Chem., 74 (2002) 3221. 

Gangl et al. [74] reported a 100-fold improvement in the detection of in vivo formed DNA adducts derived from the food-derived 2-amino-3-methyhmidaz[4,5-/ quinohne (TQ, one of the heterocyclic aromatic amines, Ch. 14.5) by the apphcation of capillary LC in combination with micro-ESI-MS. As a result, the detection limit approaches 1 adduct in 10 nucleobases using 500 pg DNA. In a subsequent study [75], this technology was applied to the quantitative analysis of the IQ-dG adduct in rat liver samples in a dose-response study. The major adduct (C8-lQ-dG) could be detected at 17.5 fmol in 300 pg of liver DNA (corresponding to 2 adducts in 10 nucleobases). 

Capillary electrophoresis (CE) MS is another technique used to separate and measure the m/z ratios of a mixture of peptides and proteins. In this method, the peptide mixture is separated by different migration rates through the electrophoresis media and the effluent is again directly sprayed into the mass spectrometer using a micro ESI device. This method is capable of femtomole or lower detection limits and is further discussed in a review. In 1996, a group at the University of Washington described a solid phase extraction (SPE) capillary electrophoresis MS-MS approach for the analysis of peptides and showed limits of detection at the 100 s of attomole level. " ... 

Nano- and micro-ESI sources that operate in the 50-300 nL/min flow range are an ideal match for typical EOF values generated on the chip (i.e., <300-400 nL/min). The dead volumes associated with nano-ESI sources are minimal. In addition, nano-ESI sources have reasonable tolerance toward buffer composition and concentration, and facilitate sensitive analysis of trace level components (even of unseparated mixtures). Moreover, due to low analyte consumption characteristics, nano-ESI analysis enables extended infusion-MS investigations, increased signal averaging capabilities. 

Martin, S.E., Shabanowitz, J Hunt, D.F, Marto, J.A. (2000) Subfemtomole MS and MS/MS Peptide Sequence Analysis Using Nano-HPLC Micro-ESI Fourier Transform Ion Cyclotron Resonance Mass Spectrometry. Anal. Chem. 72 4266-4274. 

Klein et al. used a custom-made FT-ICR MS equipped with a 22-cm-diameter horizontal bore 9.4T magnet for mass analysis. A fused-silica micro-ESI needle was used to produce positive and negative ions. Samples were infused with a flow of 400 nL/min, a needle voltage of 2 kV, and a tube lens voltage of 350 V for positive (-i-)... 

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