Fluorescence detection multidimensional approaches

Warner and co-workers justify the elaborate optical/detection system and the time commitment required per analysis on the basis of the additional sensitivity available using fluorescence detection, and on the multidimensional CD information available. For applications in which two, or more CD active fluorophores may be present, the ability to provide both an excitation and emission FDCD profile for the sample may allow differentiation of the individual components without pre-separation. Replacement of the mechanical mechanism for prism movement with an electro-optical device may improve both the SNR and reduce the time required per sample. These improvements will greatly facilitate general application of this multidimensional approach to FDCD measurements. 

Relative protein quantitation is the basis of all types of differential proteome analyses. In the 2D-gel approach protein staining with either visible or fluorescent dyes provides a reliable and sensitive method to detect changes in protein expression or isoform abundance. In the multidimensional LC approach quantitation relies mostly on stable isotope labeling and ratios between light and heavy isotopomers are determined by MS or MS/MS at the peptide level. Labeling can be performed on the protein level by... 

In the present paper we describe an apparatus for recording transient emission spectra that yields data which approach the ideal multidimensional case. We emphasize in the discussion the advantages of multichannel detection for transient emission data. We also briefly compare our approach to alternative methods for recording time and wavelength resolved fluorescence data on the picosecond time scale. 

A majority of the separation-based approaches in the microfluidic chip format for protein analysis have focused on single or multidimensional separations with detection by conventional MS or optical methods (e. g. laser induced fluorescence (LIF)). Microfluidic separation chips have also been adapted for use in immunoassays. They have also been utilized for the pre-MS sample preparation followed by conventional detection by MS. A typical flow scheme of microfluidic protein separation and detection is shown in Fig. 1. 

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