Nanospray technique

There are a number of options for analyzing a peptide mixture extracted from a gel by mass spectrometry. The most widely used techniques are MALDI-TOF, preferably with delayed extraction, and ESI with a triple-quadrupole analyzer or a combination of quadrupole and TOF analyzers. The combination of ESI with an ion trap is also of great interest due to its sensitivity and the possibility of performing structural studies by MS". Downsizing of the electrospray capillary such as in the nanospray technique results in another gain in sensitivity and an extension of the measuring time, which is important to optimize fragmentation conditions. 

Warriner, R. N., Craze, A. S., Games, D. E., and Lane, S. J. (1998). Capillary electrochromato-graphy/mass spectrometry a comparison of the sensitivity of nanospray and microspray ionization techniques. Rapid Commun. Mass Spectrom. 12, 1143-1149. 

Nanospray is a miniaturized version of electrospray. In the original setup of Wilm and Mann (8) it is utilized as an off-line technique using disposable, finely drawn (1 -gm tip), metallized glass capillaries to infuse samples at 10-30 nL/min flow rates. This allows more than 50 min analysis time with just a 1-pT sample. Due to the formation of much smaller droplets and the more effective ionization, there is often no need for LC separation, since the separation is accomplished in m/z or by MS/MS. However, limited reproducibility with respect to quantification and a more complex sample preparation can be seen as drawbacks. An on-line version for hyphenation with capillary and nano-LC as well as CE (slightly modified) is now commercially available. 

NSI Nanospray ionization (NSI) is a low-flow (10-500-nL/min) ESI technique with many advantages over conventional-flow ESI ( 200 xL/min) for the analysis of drugs, metabolites, peptides, proteins, and other macromolecules. Advantages of NSI over ESI include decreased sample consumption and increased sensitivity (Wilm and Mann, 1996 Corkery et al., 2005). NSI can be used for LC-MS or direct-infusion MS analysis of molecules (Wickremsinhe et al., 2006 Ramanathan et al., 2007c). 

Several modifications of the ESI technique have been introduced, principally micro-electrospray14 and nano-electrospray15 that have the advantage of using much lower flow rates, reducing the amount of analyte needed for a mass-spectrometric analysis this is performed using adapted probed tips. When performing nanospray, the sample is loaded into a fine hollow needle with a... 

Today, the two most common LC/MS interfaces are atmospheric pressure ionization interfaces, electrospray (ESI) and ion spray (ISI). Electrospray (Fig. 15.8) and its subtype, nanospray, are recommended for use with proteins and highly polar or ionized compounds. They are very soft ionization, concentration-dependent techniques that result in very little fragmentation and often produce multiply charged molecular ions. 

Based on this concentration dependence, modifications of the technique, called microelectrospray (pESI), or nanospray (nESI), which use much lower flow rates down to some tens of nanolitres per minute, have been developed using adapted probe tips [71-73]. Detection limits in the range of attomoles (10 15 moles) injected have been demonstrated. 

The nano-electrospray (nanoES) source is essentially a miniaturized version of the ES source. This technique allows very small amounts of sample to be ionized efficiently at nanoliters per minute flow rates and it involves loading sample volumes of 1-2 pi into a gold-coated capillary needle, which is introduced to the ion source. Alternatively for on-line nanoLC-MS experiments the end of the nanoLC column serves as the nanospray needle. The nanoES source disperses the liquid analyte entirely by electrostatic means [27] and does not require assistance such as solvent pumps or nebulizing gas. This improves sample desolvation and ionization and sample loading can be made to last 30 minutes or more. Also, the creation of nanodroplets means a high surface area to volume ratio allowing the efficient use of the sample without losses. Additionally, the introduction of the Z-spray ion source on some instruments has enabled an increase in sensitivity. In a Z-spray ion source, the analyte ions follow a Z-shaped trajectory between the inlet tube to the final skimmer which differs from the linear trajectory of a conventional inlet. This allows ions to be diverted from neutral molecules such as solvents and buffers, resulting in enhanced sensitivity. 

Van Pelt, C.K. Zhang, S. Henion, J.D. Characterizatioin of aFully Automated Nanospray System with Mass Spectrometric Detection for Proteomic Analyses, J. Biomolecular Techniques 13, 72-84 (2002). 

Nanospray ionization is a variation of regular ESI in that the typical flow rate is reduced to between 30 and 200 nL/min. Because of the very low flow rate at which samples are consumed, this technique is rapidly being integrated in many analytical applications including proteomics, metabolite characterization, and pharmaceutical analysis. An approach of combining fraction collection with automated chip-based NSI MS was recently introduced for metabolite identification (Hop, 2006 Staack et al., 2005). LC effluent was collected into a 96-well plate and the fraetions of interest were infused using an automated chip-based nanospray system for structure elucidation. 

Xie et al. [5] used vapor-deposited parylene-C to fabricate ESI tips on silicon microfluidic devices, enabling integrated liquid chromatography with mass spectrometry detection with comparable performance to conventional techniques. The drawback for these devices is the complexity involved in their fabrication, requiring many sequential photolithography steps in a clean room. However, parylene is a material with high chemical resistance and may be a useful choice for the construction of nanospray tips in future work. For example, Kameoka et al. [6] constmcted a nanospray tip comprising a parylene film sandwiched between two plastic plates (Fig. 2b). This device is relatively easy to... 

Two factors favor the use of nanoelectrospray ionization for coupling microfluidic devices to mass spectrometers. The first is the similarity between the conventional puUed-glass capillary tips and the nanospray nozzles developed for microdevices discussed above the second springs from the linear geometry of microfluidic channels. Thus, nanospray ionization techniques are probably the most likely to be used for the construction of robust interfaces between... 

A droplet evolution scheme is shown in Figure 1.5. It deals with droplets produced by nanoelectrospray. Nanospray (see Section 1.2.4) is a technique that considerably... 

Amster s apparatus is an on-line ESI-MS technique for the study of photochemical reactions that greatly reduces the transit time of photogenerated species [34]. Figure 5.5 shows sample solutions that are irradiated directly in the optically transparent nanospray tip of the ESI source. Subsequent thermal reactions of the primary photoproducts take place in the region between the photolysis zone and the tip end. The transit time of a photoproduct depends on the volumetric flow rate of the sample, the inner diameter of the tip, and D, the distance between the midpoint of the irradiated zone and the tip end. For example, with D=0.84 mm, a tip diameter of 40 im, and a flow rate of 40 tLh , products require 95 ms to arrive at the tip end for spraying. All chemical reactions are quickly quenched (ms time scale) once the... 

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