Subcellular analysis detection methods

UV absorbance is the most common detection method for any CE system, primarily because nearly all compounds absorb somewhere in the UV region of the spectrum, and also because fused-silica capillaries are transparent to UV radiation. Given these advantages, UV absorbance detectors from HPLC instruments were naturally adopted into CE systems, but a problem arises when these detectors are used for subcellular analysis subcellular analyses commonly quantify analytes that are below the LOD for UV absorbance. According to Beer s law, the sensitivity of a UV absorbance detector is related to the pathlength of the light. Therefore, UV absorbance has limited applicability to traditional CE separations due to the short path of the typical capillary diameters. 

Lastly, the most popular detection technique for subcellular analysis is LIF due to the extremely low LODs that can be attained. Mass LODs on the order of attomoles can be achieved with commercial CE-LIF instruments and custom-built instruments, in conjunction with sheath-flow cuvettes, can reach down to the yoctomole (10 " mole) scale. LIF detection thus allows the quantification of minute amounts of analyte, and is the only detection method that has been used to detect individual organelles. We will therefore describe off-column LIF detection in further detail. 

Cellular Metabolites. - A review of methods for the measurement of ml has been produced with 95 references. It examines the quantitative measurement of ml by mass spectrometry and in vivo NMR. The NMR chemical shifts and /-coupling values of 35 metabolites which can be detected by in vivo or in vitro investigations of the mammalian brain have been published. The principles and recent applications of dynamic nuclear polarisation, which combines the sensitivity to oxygen of EPR and the tractability of NMR imaging, have been reviewed with 244 references. A review of studies of intermediary metabolism, including the use of NMR in the analysis of substrate selection under in vivo conditions, has been produced. A review has been produced, with 74 references, on the study of metabolic flux and subcellular transport of metabolites using NMR. " ... 

Objective analysis of the composition of the phases developing on-line measurement technique fermentor-interface, instrument (computer) Difficulty great number + variety of components/selective analytical method not sensitive to interferences separation before detecting sterile conditions (built-in sensors, airtight scaling, endurance of strerilization) on subcellular level biochemical informations are limited... 

Cells and tissues must be lysed to release the antigen protein for analysis by immunoblotting and immunoprecipitation. The method of extraction depends upon the subcellular localization of the antigen and the subsequent mode of analysis and detection. The methods described in this section are intended as guides, and can be adapted to optimize the extraction of individual proteins. 

---