Clinical Chemistry
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Clinical Chemistry 35: 2082-2086, 1989;
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Clinical Chemistry, Vol 35, 2082-2086, Copyright © 1989 by American Association for Clinical Chemistry

Mechanisms responsible for increasing immunoreactivity of apolipoprotein A-I with storage: the role of oxidation

XL Wang, NP Dudman, J Wang and DE Wilcken
Department of Cardiovascular Medicine, Prince Henry Hospital, University of New South Wales, Sydney, Australia.

Atherogenesis has been linked to low concentrations of high-density lipoprotein and its principal carrier protein, apolipoprotein (apo) A- I, in serum. We measured apo A-I by enzyme-linked immunosorbent assay, using polyclonal antiserum raised against purified apo A-I and conjugated to peroxidase. During storage at 4 degrees C and above, the immunoreactivity of apo A-I increased so that measured values in stored serum were difficult to interpret. The immunoreactivity of serum apo A- I also increased substantially (up to 50-fold) during exposure to NaIO4. This increase was apo A-I concentration dependent, was linearly dependent on the concentration of NaIO4 up to 200 mmol/L at pH 7.4 and 25 degrees C, and was not ascribable to nonspecific binding of antibody- peroxidase conjugate to the reaction wells. Treatment with NaIO4 caused apo A-I immunoreactivity to increase to a peak or plateau, then gradually decline. In 100 mmol/L NaIO4 (pH 7.4, 25 degrees C), this plateau occurred 30-60 min after oxidation commenced. When frozen serum and stored nonfrozen serum were both oxidized with NaIO4 under the same conditions, the apo A-I from the frozen serum had a significantly (P = 0.0004) greater increase in immunoreactivity. Sera from different patients with different apo A-I concentrations did not have proportionately increased immunoreactivity after treatment with NaIO4. Our findings suggest that the increase in apo A-I immunoreactivity on storage at 4 degrees C or higher involves atmospheric oxidation, and they raise the possibility of in vivo oxidized and nonoxidized forms of apo A-I.


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Copyright © 1989 by the American Association for Clinical Chemistry.