Journal
NATURE BIOTECHNOLOGY
Volume 19, Issue 10, Pages 946-951Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/nbt1001-946
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Funding
- NCI NIH HHS [1R33CA84698] Funding Source: Medline
- NHLBI NIH HHS [R01 HL067569, R01 HL067569-02, R01 HL067569-01, HL67569] Funding Source: Medline
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An approach to the systematic identification and quantification of the proteins contained in the microsomal fraction of cells is described. It consists of three steps: (1) preparation of microsomal fractions from cells or tissues representing different states; (2) covalent tagging of the proteins with isotope-coded affinity tag (ICAT) reagents followed by proteolysis of the combined labeled protein samples; and (3) isolation, identification, and quantification of the tagged peptides by multidimensional chromatography, automated tandem mass spectrometry, and computational analysis of the obtained data. The method was used to identify and determine the ratios of abundance of each of 491 proteins contained in the microsomal fractions of naive and in vitro-differentiated human myeloid leukemia (HL-60) cells. The method and the new software tools to support it are well suited to the large-scale, quantitative analysis of membrane proteins and other classes of proteins that have been refractory to standard proteomics technology.
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