Determination of advanced oxidation protein products (AOPP)
We determined an oxidate stress biomarker, advanced oxidation protein products (AOPP)
concentrations in patients in Department of Cardiovascular Surgery, Semmelweis University,
Budapest. Blood was drawn from the median cubital vein. Plasma samples were prepared
from EDTA-anticoagulated blood after centrifugation at 3000 rpm (1660 g) for 10 min and
were stored at -20ºC until assay.
AOPP are measured by spectrophotometry at 340nm under acidic conditions. The spectral
characteristics of AOPP correspond to several chromophores which include dityrosine,
carbonyls and pentosidine but not nitrotyrosine. The AOPP formation is calibrated by the
formation of tri-iodide ion upon oxidation of potassium iodide with chloramine-T. The triiodide
ion absorbance at 340nm gives a linear calibration curve within the range of 0–100
mmol/l of chloramine-T. AOPP concentrations are expressed in micromoles per litre of
chloramine-T equivalents.
Here we describe an automated version of the originally published technique that we adapted
for a Cobas Mira Plus (Roche, Basel, Switzerland) clinical chemistry analyzer. A 40-mL
aliquot of plasma sample was added to 160 mL of phosphate-buffered saline (PBS) reagent,
mixed and incubated for 25 s and then the absorbance of the mixture was read at 340 nm.
Then 20 mL of acetic acid was added, mixed and incubated for 25 s. Next, 10 mL of
Potassium iodide (KI) solution was added to the reaction mixture, mixed and after an elapse
of 25 s the absorbance was read again at 340nm.