Freeze-thaw and matrix effects in direct high-density lipoprotein cholesterol methods.
BACKGROUND
There is frequent discussion on the susceptibility of direct high-density lipoprotein cholesterol (HDL-C) methods to matrix effects. In Vitro Diagnostics manufacturers recognize this issue and regularly improve their HDL-C reagent formulations in subsequent generations.


METHODS
The 3rd generation direct HDL-C assay from Roche was investigated for matrix effects in comparison to the former generation, a Beckman direct HDL-C method and a conventional phosphotungstate (PTA)/Mg(2+) precipitation method. In addition, 235 heparin plasma samples were measured freshly and after a freeze-thaw cycle with the Roche 2nd and 3rd generation direct HDL-C. Biases, outliers, and intraclass correlation coefficients (ICCs) were calculated for both experiments. Multivariate analysis was used to investigate interference by matrix components.


RESULTS
In fresh samples, Roche 2nd and 3rd generation HDL-C methods averaged +0.15 mmol/L (95% CI: 0.13-0.16) and +0.08 mmol/L (95% CI: 0.07-0.09) higher compared to frozen samples. In frozen aliquots, ICCs for Roche 2nd and 3rd generation and Beckman direct HDL-C as compared to PTA/MgCl(2) were 0.963, 0.966, and 0.924, respectively. Predictors of outliers (defined as having an absolute difference >0.21 mmol/L) in comparisons of direct methods to the PTA/MgCl(2) precipitation method were high triglyceride and low albumin levels.


CONCLUSIONS
The 3rd generation direct HDL-C from Roche has become insensitive to most matrix effects, bringing along more accurate results in hypoalbuminemic and hypertriglyceridemic samples. Surprisingly, Roche direct assays produced significantly higher HDL-C levels in fresh samples compared to frozen plasma samples. If confirmed by others, the latter finding has implications for patient management and necessitates further reagent optimization.
Cobbaert
Christa
C
Department of Clinical Chemistry and Hematology, Amphia Hospital, Breda, The Netherlands. ccobbaert@amphia.nl
van Haperen
Corrie
C
Bastijns
Claudia
C
Demacker
Pierre N M
PN
D001774:Blood Chemical Analysis / Q000295:instrumentation / Q000379:methods*; D008076:Cholesterol, HDL / Q000097:blood*; D005615:Freezing; D006801:Humans; D011237:Predictive Value of Tests; D012044:Regression Analysis; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D008076:Cholesterol, HDL