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Isolated subcellular fractions have been instrumental in elucidating cell function. The use of such fractions for the identification and biochemical characterization of subcellular organelles, combined with cell- free systems, has provided key insights into the function and machineries of organelles, including those involved in vesicle transport, quality control and protein sorting. Despite their obvious utility, popular cell biology has come to regard in vitro-based approaches as inferior to in vivo-based approaches. Usual criticisms are contamination, non-representative processes and an inability to recreate the dynamic processes seen in vivo. In a similar way, proteomics has been viewed with reservation. Despite this, and building on the tradition of in vitro-based approaches, organelle proteomics based on liquid chromatography and tandem mass-spectrometry has recently made significant contributions to cell biology, and now allows the molecular machineries of organelles to be defined with high precision. |
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