Redistribution of metal ions to control low density lipoprotein oxidation in Ham's F10 medium.
The study of cell-mediated low density lipoprotein (LDL) oxidation has traditionally been undertaken using Ham's F10 media due to its high metal content and low levels of antioxidants. Although there has been no acknowledged change to this media in recent years by the suppliers, Ham's F10 medium has been found to be extremely inconsistent in its promotion of LDL oxidation in the absence of cells. This variability contrasts with the relatively consistent rates of THP-1 cell-mediated LDL oxidation. This study has now shown that the variability in cell-free LDL oxidation is medium-dependent and not an artefact of experimental protocol. It presents evidence that suggests the variable rates of cell-free LDL oxidation are caused by iron auto-oxidation during storage of the Ham's F10 medium. The medium can be standardized by removal of all transition metals, by treatment with Chelex, before the addition of known amounts of iron or copper. This treatment generates a cell culture medium that only allows very slow LDL oxidation in the absence of cells.
Firth
Carole A
CA
Free Radical Biochemistry Laboratory, School of Biological Sciences, University of Canterbury,Private Bag 4800, Christchurch, New Zealand.
Gieseg
Steven P
SP
D002460:Cell Line; D002474:Cell-Free System; D003300:Copper / Q000378:metabolism; D003470:Culture Media / Q000378:metabolism*; D006801:Humans; D007477:Ions; D007501:Iron / Q000378:metabolism; D007552:Isotonic Solutions / Q000737:chemistry*; D008077:Lipoproteins, LDL / Q000737:chemistry / Q000378:metabolism*; D008956:Models, Chemical; D010100:Oxygen / Q000737:chemistry / Q000378:metabolism*; D013438:Sulfhydryl Compounds; D017392:Thiobarbituric Acid Reactive Substances / Q000737:chemistry; D013997:Time Factors; D024502:alpha-Tocopherol / Q000737:chemistry
D003470:Culture Media; D007477:Ions; D007552:Isotonic Solutions; D008077:Lipoproteins, LDL; D013438:Sulfhydryl Compounds; D017392:Thiobarbituric Acid Reactive Substances; C083923:nutrient mixture F-10 Ham; D003300:Copper; D007501:Iron; D024502:alpha-Tocopherol; D010100:Oxygen