Analytical variables affecting exchangeable copper determination in blood plasma

Abstract

To resolve discrepancies observed in the determination of plasma exchangeable Cu (also called direct reacting Cu or loosely bound Cu) by several methods, plasma storage techniques and various aspects of a stable isotope dilution procedure for exchangeable Cu were evaluated. Results indicated that the exchangeable Cu fraction of plasma increased with storage at room temperature, at 5 degrees C and when subjected to repeated freeze/thaw cycles. Samples could be safely stored at -65 degrees C. Exchange between added 65Cu2+ and endogenous plasma Cu rapidly went to completion in the isotope dilution procedure. Analytical results were unaffected by shaking method, sample size or the presence of heparin. A small difference was observed between serum and plasma. The determination of exchangeable Cu did not vary over a period of 4 h when plasma was exposed to 1.6 x 10(-4)-M sodium diethyldithiocarbamate (used in the isotope dilution method) but steadily increased when exposed to 1.1 x 10(-2)-M sodium diethyldithiocarbamate, which suggested that tightly bound Cu (probably in ceruloplasmin) was exchanging with isotopic tracer at the higher concentration. Determination of exchangeable Cu was constant from pH 7.2-8.5 but increased substantially at higher pH. Complete recovery of natural Cu added to plasma was obtained. Studies in solution indicated that 65Cu2+ exchanged readily with albumin- and amino acid-bound Cu. Ultrafiltration of plasma yielded a Cu fraction about half that of the exchangeable Cu fraction. We conclude that the stable isotope dilution procedure for plasma exchangeable Cu yields reliable, physiologically meaningful results.

Relative exchangeable copper: A valuable tool for the diagnosis of Wilson disease

“…Exchangeable copper corresponds to the labile fraction of copper complexed within plasma proteins (such as albumin, transcuprein) and ultrafiltrable copper representing copper bound to low molar mass molecules (such as amino acids) but not complexed within ceruloplasmin . REC has been validated in a first study including 16 WD patients, mostly neurological forms, 25 wild‐type homozygous and 45 heterozygous stemming from familial screening and 62 healthy subjects: a REC>18.5% was an excellent discriminatory tool for the diagnosis of WD offering 100% sensitivity and 100% specificity in this population, what makes it in theory the ideal tool for diagnosis.…”

Biophysical Studies of the Induced Dimerization of Human VEGF Receptor 1 Binding Domain by Divalent Metals Competing with VEGF-A

“…They are the second and the third most abundant trace metals in plasma (16.5 ± 8.6 μM and 16.6 ± 6.2 μM, respectively) [42]. Plasmatic zinc is bound to and transported by albumin and transferrin, and the stock is rapidly exchangeable [43], while the major part of plasmatic copper is tightly bound to ceruloplasmin, with a low exchangeable fraction (0.5 μM) [44]. Consequently, the IC 50 that we observed in our displacement assays are on the same order of magnitude than exchangeable Cu 2+ and Zn 2+ concentrations in plasma.…”

Evolution of Exchangeable Copper and Relative Exchangeable Copper through the Course of Wilson's Disease in the Long Evans Cinnamon Rat

“…This leads to an elevation of urinary copper. CuEXC has been described as the labile fraction of copper bound to different molecules such as albumin and transcuprein [8]. Hence, an increase in CuEXC serum levels could indicate the presence of abnormally high amount of toxic extrahepatic copper.…”

“…In that study, REC provided 100% specificity and sensitivity. CuEXC corresponds to the labile fraction of copper bond mainly to albumin [7], [8], [9]. An increase of CuEXC superior to normal levels is thought to reflect a blood and tissue copper overload that occurs after saturation of hepatocytes and the spillage of Cp unbound copper into the blood owing to hepatic cytolysis.…”