Evaluation of the Abbott IMx fluorescence polarization immunoassay and the bio-rad enzyme immunoassay for homocysteine: comparison with high-performance liquid chromatography.
We evaluated the precision, linearity and accuracy of the Abbott IMx and Bio-Rad (Axis) homocysteine assays. Both assays make use of S-adenosyl-homocysteine hydrolase and excess adenosine, to convert homocysteine to S-adenosylhomocysteine (SAH). A monoclonal anti-SAH antibody is then used to quantify SAH. The IMx assay measures the fluorescence polarization of a conjugated SAH analogue for the final analytical step, whereas the Bio-Rad method uses a microplate enzyme immunoassay (EIA) employing an anti-mouse antibody peroxidase conjugate. The Abbott procedure is completely automated whereas the Bio-Rad EIA is performed manually. Between-run coefficient of variation using commercial controls was 2.6% at 7 micromol/L, 2.5% at 13 micromol/L and 1.7% at 24 micromol/L for the Abbott method, and 19.7% at 6.4 micromol/L, 15.9% at 11.0 micromol/L and 14.5% at 23.4 micromol/L for the Bio-Rad method. Both assays correlated well with a high-performance liquid chromatography (HPLC) procedure for homocysteine: Bio-Rad EIA = 1.03HPLC + 1.0 micromol/L, r=0.98, s(y/x)=0.51; Abbott IMx = 1.02HPLC + 0.7 micromol/L, r=0.99, s(y/x) = 0.33. Both methods were linear up to 50 micromol/L homocysteine. The IMx assay had superior precision as well as the technological advantage of being completely automated. Both immunoassays exhibited greatly improved throughput compared with our existing HPLC method.
Donnelly
J G
JG
Department of Laboratory Medicine, Ottawa Hospital, Ontario, Canada. jdonnelly@civich.ottawa.on.ca
Pronovost
C
C
D002851:Chromatography, High Pressure Liquid; D005454:Fluorescence Polarization; D006710:Homocysteine / Q000097:blood*; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D011933:Reagent Kits, Diagnostic; D012044:Regression Analysis; D015203:Reproducibility of Results; D012435:S-Adenosylhomocysteine / Q000276:immunology / Q000378:metabolism; D012680:Sensitivity and Specificity
D011933:Reagent Kits, Diagnostic; D006710:Homocysteine; D012435:S-Adenosylhomocysteine