High Molecular Weight Analytes

TABLE Suggested IP Values for the Ideutiticatiou of Peptides and Proteius Smaller Thau 8kDa Usiug Mass Spectrometry aud Immuuoafiiuity Purificatiou  

Low resolution MS precursor ion/molecular weight detenninatiou Low resolution MS product ion 

High resolution/high accuracy MS product ion Immunoaffinity purification 

2 MODERN MASS SPECTROMETERS IN DOPING CONTROLS ADVANTAGES AND DISADVANTAGES OF AVAILABLE TECHNIQUES 

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The dissolution of high molecular weight analytes increases the viscosity of the samples. Samples with a high viscosity can exhibit viscous fingering on the column top. Because this phenomenon can impair resolution, it is recommended to keep the sample concentration below certain limits. These limits are shown in Table 11.5. 

A further extension of the DFG S19 method was achieved when polar analytes and those unsuitable for GC were determined by LC/MS or more preferably by liquid chromatography/tandem mass spectrometry (LC/MS/MS). Triple-quadrupole MS/MS and ion trap MS" have become more affordable and acceptable in the recent past. These techniques provide multiple analyte methods by employing modes such as time segments, scan events or multiple injections. By improving the selectivity and sensitivity of detection after HPLC separation, the DFG S19 extraction and cleanup scheme can be applied to polar or high molecular weight analytes, and cleanup steps such as Si02 fractionation or even GPC become unnecessary. 

Guo XQ, Castellano FN, Li L, Lakowicz JR (1998) Use of a long lifetime Re(I) complex in fluorescence polarization immunoassays of high-molecular weight analytes. Anal Chem 70 632-637... 

Cyanines Fluorescent Labels for Low- and High-Molecular-Weight Analytes... 

The synthesis, spectral properties, and applications of symmetrical as well as unsymmetrical, hydrophobic oxo-squaraine probes for noncovalent interaction with proteins, lipids, cells, and other high-molecular-weight analytes are described in numerous publications and patents [52, 57, 58]. 

The introduction of soft ionization techniques, such as plasma desorption (PD),[1] field desorption (FD)[2] and fast atom bombardment (FAB),[3] marked the beginning of a new era for MS. In fact, they allowed MS to extend its applications to wide classes of nonvolatile, polar, thermally unstable and high molecular weight analytes. This opened up new horizons for MS in many unexpected fields, such as biology, biomedicine and biotechnology, in which this methodology had not previously found any possible application. 

The mechanism governing the analysis of high molecular weight analytes GPC. 

Employing variations of the ELISA technique, the fluorogenic ELISA assays are based on the capture of antigens or antibodies from a solution by an insoluble solid phase. These assays are used to determine concentrations of high molecular weight analytes, to indicate the presence of specific antibodies, or for competitive analysis of low-concentration haptens. A linear capillary... 

The slow mass transfer of polymeric packings is limited to small molecules. High-molecular-weight analytes such as proteins or synthetic polymers do not suffer from this problem, which is one of the reasons why the predominant application of polymeric packings is in the chromatography of macromolecules. 

There have been several new developments in fluorescence polarization immunoassay (FPIA). Lackowicz and Terpetschnig reviewed the use of long-lifetime metal-ligand complexes in fluorescence polarization assays [154, 155]. New complexes with Re(l) and Ru(II) were described for the highly sensitive detection of high-molecular weight analytes by FPIA. Laser-induced fluorescence polarisation detection has been used by Yatscoff and coworkers in capillary electrophoresis detection (CE-LIFP) [156]. For the analyte cyclosporin picomolar detection limits were attained. 

Common interfaces include the molecular jet and flow spHtters. The first uses the difference in momentum between the low-molecular weight carrier gas and the high-molecular-weight analytes. The column effluent passes into the separator inlet line, which is enclosed in a glass chamber that is under vacuum. A small gap separates this line from the outlet hne. Sample molecules move preferentially from one line to the other by inertia, while much of the carrier gas is removed tangentially by the vacuum. The enrichment also increases the sensitivity of GC-MS. 

There are both nonpolar and polar coatings available. Dimethylpolysi-loxane is the most popular nonpolar one, and the 7 pm thin film is best for high molecular weight analytes the 30 jum film is preferable for mid-range (pesticides), and the 100 /tm film for volatiles. Extraction efficiency depends on many factors the chemical nature of the analyte, the sample matrix and the polymer coating the extraction time and temperature the degree of stirring and the analyte concentration. The desorption step depends primarily on the injection port temperatures, the volatility of the analyte, and the film thickness. 

Desorption methods are also useful for the ionization of both ionic and neutral molecules. FD, LD, FAB, and SIMS have been used for the characterization of nonvolatile metal complexes. MALDI is probably the most convenient method for analysis of high-molecular-weight analytes, such as complex biomolecules, metal-containing polymers, etc. 

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