Analytical validation of the PRO-Trac II ELISA for the determination of tacrolimus (FK506) in whole blood.
BACKGROUND
The analytical validation of multiple lots of the PRO-Trac II ELISA (DiaSorin) for the determination of tacrolimus in whole blood is described.


METHODS
The analytical parameters assessed included analytical sensitivity, dilution linearity, functional sensitivity, values in samples containing no tacrolimus, intra- and interassay precision, supplementation and recovery, metabolite cross-reactivity, interference studies, and method comparisons HPLC-tandem mass spectrometry (HPLC/MS/MS) and the IMx Tacrolimus II multiparticle enzyme immunoassay. Where appropriate, assessments were performed according to NCCLS guidelines.


RESULTS
The mean analytical detection limit was <0.25 microg/L for all lots, whereas the functional sensitivity was 1.0 microg/L. Excellent linear correlation (r = 0.985) was observed for dilution linearity. The intraassay imprecision was <7%, and the total imprecision by ANOVA was <10%. Recovery was 109% +/- 11%. Metabolite cross-reactivity was consistent with previous reports for this antibody. No interference was observed for 35 tested drugs. Method comparison with HPLC/MS/MS showed no statistically significant differences. Samples exhibited stability through four freeze/thaw cycles and for 1 week at room temperature.


CONCLUSIONS
These data demonstrate that the PRO-Trac II ELISA is a robust, accurate, and precise tool for the assessment and management of tacrolimus blood concentrations in transplant patients.
MacFarlane
G D
GD
DiaSorin Inc., Stillwater, MN 55082, USA. gordon.macfarlane@diasorin.com
Scheller
D G
DG
Ersfeld
D L
DL
Shaw
L M
LM
Venkatarmanan
R
R
Sarkozi
L
L
Mullins
R
R
Fox
B R
BR
D002851:Chromatography, High Pressure Liquid; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D007166:Immunosuppressive Agents / Q000097:blood*; D016031:Liver Transplantation; D013058:Mass Spectrometry; D011446:Prospective Studies; D011933:Reagent Kits, Diagnostic; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D016559:Tacrolimus / Q000097:blood*
D007166:Immunosuppressive Agents; D011933:Reagent Kits, Diagnostic; D016559:Tacrolimus