Biochemical characterization of air-filled albumin microspheres.
Heating and sonication of a solution of human serum albumin (HSA) yields air-filled microspheres that can be used as a contrast agent in ultrasound examinations. The microspheres are stabilized by a thin-layer of protein surrounding the air bubbles. As long as the microspheres were intact, the protein was insoluble in aqueous solutions. After disintegration of the microspheres, the protein could be solubilized in several solutions. The intermolecular interactions in the microsphere protein have been elucidated from its solubility properties. The microspheres were disintegrated by several detergents which also solubilized the protein. After pressure disintegration of the microspheres, the protein was solubilized immediately in urea and guanidinium chloride, and also in phosphate-buffered saline after incubation overnight. These results indicate that the protein was mainly stabilized by non-covalent forces. The solubilization in buffer was inhibited by a high salt concentration, suggesting that hydrophobic interactions were involved in stabilizing the microsphere structure. Analysis of the solubilized protein by gel filtration showed that the protein contained substantial amounts of soluble aggregates of HSA. Reduction of the disulphide bonds dissolved these aggregates into monomeric HSA, showing that intermolecular disulphide bonds were also involved in stabilization of the microspheres. The solubilized protein also contained less fatty acids than the soluble HSA used for the production of microspheres. These results show that the microsphere protein has the same characteristics as heat-denatured HSA.
Hellebust
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H
Nycomed Imaging AS, Oslo, Norway.
Christiansen
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C
Skotland
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T
D000388:Air; D002021:Buffers; D002850:Chromatography, Gel; D004220:Disulfides / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D006358:Hot Temperature; D006801:Humans; D007150:Immunohistochemistry; D007525:Isoelectric Focusing; D008863:Microspheres; D012709:Serum Albumin / Q000737:chemistry* / Q000502:physiology; D012995:Solubility
D002021:Buffers; D004220:Disulfides; D012709:Serum Albumin