Rapid size-exclusion chromatography of proteoglycans.
The separation of intact proteoglycans using high-performance liquid chromatography is not trivial because the high molarity denaturing buffers required to maintain proteoglycans in the disaggregated state create back-pressures higher than the limits of many HPLC systems. Until recently, low back-pressure requirements of HPLC size-exclusion columns precluded their use for the separation of intact proteoglycans. In this study we show that rapid size-exclusion chromatography is possible in 8 M urea buffers using a Dionex BioLC system equipped with a Bio-Rad BioSil Sec-400 column. This technique reduced the time required for size-exclusion chromatography of intact proteoglycans from approximately 18 h (Sepharose CL4B) to 25 min and in some cases improved resolution of the sample.
Johnson
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Department of Biochemistry, Meharry Medical College, Nashville, TN 37208, USA.
Blake
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D000818:Animals; D002813:Chondrosarcoma; D002847:Chromatography, Agarose; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002852:Chromatography, Ion Exchange; D011509:Proteoglycans / Q000032:analysis* / Q000096:biosynthesis; D051381:Rats; D013431:Sulfates / Q000032:analysis / Q000378:metabolism; D013462:Sulfur Radioisotopes; D013997:Time Factors; D014407:Tumor Cells, Cultured
D011509:Proteoglycans; D013431:Sulfates; D013462:Sulfur Radioisotopes; C012036:sodium sulfate