Analytical & pre-analytical variabilityAnalytical & pre-analytical variability

FibroTest and FibroMax

In this presentationIn this presentation

Pre-analytical variabilityPre-analytical variability

1.1. Fasting versus non-fastingFasting versus non-fasting

2.2. Transport, pre treatment and storageTransport, pre treatment and storage

3.3. USA specifications for FibroSure (USA trade mark for FibroTest)USA specifications for FibroSure (USA trade mark for FibroTest)

Analytical variabilityAnalytical variability

4.4. Inter-laboratory variabilityInter-laboratory variability

5.5. Intra-laboratory variabilityIntra-laboratory variability

6.6. Transferability between analyzersTransferability between analyzers

Pre-analytical variabilityPre-analytical variability

Fasting, Storage, US specifications

Fasting versus non fastingFasting versus non fastingMunteanu et al, Comp Hep 2004Munteanu et al, Comp Hep 2004

• Intra-individual fasting versus postprandial variation of biochemical markers of liver fibrosis (FibroTest, FT) and (ActiTest, AT)

Patients and methods

64 subjects

Measurement of variation +/- 1 hour from meal

Results No significant impact on FT/AT components

ANOVA >0.09 and coefficient of variation (CV) from 0,09 for FT to 0,13 for AT Spearman correlation coefficient from 0,87 to 0,995

Significant impact on triglycerides and glucose ANOVA P=0.01, CV= 0.65 and ANOVA P=0.04, CV=0.31

Concordance between fasting and predicted histological stages and grades FT: kappa=0.91, AT: kappa=0.80 almost perfect

Conclusion For FibroTest: Fasting is not a requirement for to perform FibroTest, which make the screening for patients at risk of liver diseases more convenient

For FibroMax: Due metabolic components of the test, fasting is required to perform a valid FibroMax (especially NashTest and SteatoTest

Transport, pre treatment and storageTransport, pre treatment and storage- Messous et al, Clinica Chimica 2005- Messous et al, Clinica Chimica 2005

• Time impact of serum storage temperature on stability of important liver enzymes and proteins and on FibroTest-ActiTest

Conclusions

FibroTest ActiTest results diminish more than

fifty percent with -20°C and -35°C storage.

Sera storage at –80°CSera storage at –80°C is recommended for retrospective analyses.

To avoid the risk of false negative or false positive results for non-invasive liver markers like FibroTest-ActiTest, we recommend validation studies on fresh sera.

ActiTest

USA specifications for FibroSure USA specifications for FibroSure (USA trade mark for FibroTest)(USA trade mark for FibroTest)

FIBROSURE™ 550123

CPT Pending

Synonyms HCV FIBROSURE™; FIBROSURE™, FibroTest, ActiTest

Test Includes FibroTest (alpha2-macroglobulin, haptoglobin, apolipoprotein A1, bilirubin, gamma glutamyl transpeptidase [GGT]);

ActiTest (alpha2-macroglobulin, haptoglobin, apolipoprotein A1, bilirubin, gamma glutamyl transpeptidase [GGT], alanin aminotransferases [ALT])

Specimen Serum Volume 4 mL Minimum Volume 2 mL (Note: This volume does not allow for repeat testing.)

Container Red-stopper tube or serum-separator tube

Collection Separate serum and refrigerate at 2°C to 8°C. Specimen is stable for as long as three days. Freeze if

Storage longer than 72 hours is needed.

Storage Instructions Refrigerate –80°C

USA specifications for FibroSure USA specifications for FibroSure (USA trade mark for FibroTest)(USA trade mark for FibroTest)

Reference Interval METAVIR scale

Fibrosis stageF0: no fibrosisF1: portal fibrosisF2: bridging fibrosis with few septaF3: bridging fibrosis with many septaF4: cirrhosis

Necroinflammatory activity gradeA0: no activityA1: minimal activityA2: moderate activityA3: severe activity

Use noninvasive measure

of fibrosis and necroinflammatory activity in the liver of individuals with chronic hepatitis C virus

Limitations Because this procedure is new, Medicare and other carriers may not yet recognize it as a covered benefit for patients.

Methodology Patented Artificial Intelligence algorithm combines patient’s age, gender, and the results of six biomarkers to generate a measure of fibrosis and necroinflammatory activity in the liver

Analytical variabilityAnalytical variability

Fasting, Storage, US specifications

Inter-laboratory VariablityInter-laboratory VariablityHalfon et al, Comp Hep 2002Halfon et al, Comp Hep 2002

• A prospective assessment of the inter-laboratory variability of biochemical markers of fibrosis (FibroTest) and activity (ActiTest) in patients with chronic liver disease

Concordance (kappa coefficient) between laboratories and the reference center.

FibroTest calculated with GGT expressed in International Unit ActiTest calculated with GGT-ALT, expressed in International Units

ConclusionsConclusions Standardized methods against a reference method , as well as calibrated analyzers should be used to increase

inter-laboratory coherence, particularly for GGT and ALT measurements. ALT and GGT expressed as multiples of the upper limit of normal should be avoided.

Intra-laboratory variabiltyIntra-laboratory variabilty- Imbert-Bismut et al, Clin Chem Lab Med 2004- Imbert-Bismut et al, Clin Chem Lab Med 2004

• Intra-laboratory analytical variability of biochemical markers of fibrosis (FibroTest) and activity (ActiTest) and reference ranges in healthy blood donors.

Total imprecision for FT-AT and its components on pooled sera (NCCLS-EP5-T)

The total imprecision is correct for the different parameters. A VC of 8.8% for total bilirubin was acceptable considering that this variation occurred for the low mean values

(mean 7.5 micromoles/L).

Intra-laboratory variabiltyIntra-laboratory variabilty- Imbert-Bismut et al, Clin Chem Lab Med 2004- Imbert-Bismut et al, Clin Chem Lab Med 2004

Intra-patient reproducibility of components and of FibroTest-ActiTest

Concordance between two consecutive measurements was strong (kappa=0.88) for cirrhosis (F4) versus non-cirrhosis.

Stages F0/F1, F2 and F3F4 had kappa coefficients of 0.83 and the concordance was moderate for the repeated ActiTests (kappa=0.44)

Intra-laboratory variabiltyIntra-laboratory variabilty- Imbert-Bismut et al, Clin Chem Lab Med 2004- Imbert-Bismut et al, Clin Chem Lab Med 2004

ConclusionsConclusions

The reproducibility of two FibroTests in the same patient evaluated four days apart is good despite some isolated variations of the components. This however, has no impact due to the linearization of the results.

The homogeneity of the results could be attributed to the homogeneity of adapted techniques on the different analyzers (standardized techniques compared to reference techniques), as well as the same conditions of calibration of the enzymatic activity measurements.

The study demonstrated the analytical reliability of FibroTest and ActiTest and the importance of the continued development of homogeneity and good transferability of the results between analyzers and laboratories.

Transferability between analysersTransferability between analysers- Imbert-Bismut et al, Ann Biol Clin 2005- Imbert-Bismut et al, Ann Biol Clin 2005

• Results transferability on RXL, ARX, X-Pand, BN2 (Dade Behring) and modular DP (Roche Diagnostics) analyzers: application to component assays of FibroTest and ActiTest (on 150 serum samples)

Methods of measurement adapted on different automates and evaluation methods

Transferability between analysersTransferability between analysers- Imbert-Bismut et al, Ann Biol Clin 2005- Imbert-Bismut et al, Ann Biol Clin 2005

Results of inter-platform comparisons of Dade Behring: RXL, ARX, X-Pand and results ofBN2-Modular / RXL: regression models according to Passing- Bablok.

ConclusionsConclusions This study showed harmonious results for each parameter between the Dimension analysers and

between RXL and BN2- Modular DP. Disregarding alpha-2 macroglobulin, which cannot be assayed on RXL, the results of FibroTest

and ActiTest were similar to those performed on BN2- Modular DP and RXL.

Transferability between analyzersTransferability between analyzers

All validated analyzers and reagents are listed in the technical recommendations available on www.oneliver.com in the

“For laboratories” section