current practices in inr testing in belgian …...commissie voor klinische biologie commission de...

Commissie voor Klinische BiologieCommission de Biologie Clinique

Current practices in INR testing in Belgian laboratories

Marjan Van Blerk, WIV-ISP, Brussels

Rue Juliette Wytsmanstraat 14 | 1050 Brussels | BelgiumT +32 2 642 51 11 | F +32 2 642 50 01 | email: info@iph.fgov.be | www.iph.fgov.be

• Belgian EQAS in Coagulation

• Survey 2008/3

Belgian EQAS in Coagulation

• 3 surveys per year• 3 lyophilized plasmas per survey screened

for hep B, hep C, syphilis and HIV 1 & 2• For PT testing:

- artificially depleted plasmas - pools of plasma samples from patients

treated with oral anticoagulants • Participation is mandatory

Belgian EQAS in Coagulation

• Statistical evaluation by a non-parametricapproach: method of Tukey

• Report of individual results• Unacceptable result:

outside ± 2 SD of method median value• Discussion of results within the advisory board• Global report

Evolution of overall CV, %

12.7 (n=223)

10.5 (n=329)3.5 – 4

9.7/10.4(n=223)

8.4 (n=329)2.5

20061997Median INR value

Survey 2006/3

• to assess the practices in INR testing in the Belgian and Luxembourg laboratories

• to determine the extent to whichrecommendations and guidelines are followed

Survey 2006/3

2 pools of plasma samples from patients on oral anticoagulants

1 artificially depleted plasma

Questionnaire

Survey 2006/3: Questionnaire

• Citrate concentration• Lot number thromboplastin• Calculation vs calibration of INR• If calculation:

ISI: Manufacturer/Local calibrationNormal PT: MNPT/Manufacturer/

Pool/Lyophilized plasma…• Validation of INR

Survey 2006/3: INR results

22412.73.8515.73.079.72.29

NCV,%INR 7400

CV,%INR7399

CV,%INR7395

Survey 2006/3: ConclusionsNeed for further standardization of PT testing in Belgium

Recommended to•use 0.105-0.109 mol/L trisodium citrate as anticoagulant•use homogeneous test systems (reagent/instrument)•determine the system-specific MNPT

Cave: local ISI calibration

Validation of INR: Need for guidelines and control materials

Evolution of overall CV, %

7.63.132008/26.63.152008/18.22.092007/39.43.792007/2

CV, %Median INR valueSurvey

Survey 2008/3

Repeat the study of 2006/3 to evaluate:

• whether the tendency to decrease of the overall CV could be confirmed

• and if so, if this could be related to changes in methodology

Survey 2008/3: Samples

3 pools of plasma samples frompatients on oral anticoagulants

CO/8610CO/8883CO/8884

Survey 2008/3: Questionnaire

• Citrate concentration• Lot number thromboplastin• Calculation vs calibration of INR• If calculation:

ISI: Manufacturer/Local calibrationNormal PT: MNPT/Manufacturer/

Pool/Lyophilized plasma…• Validation of INR

Survey 2008/3: Participation

Survey202 Belgian and 14 Luxembourg participants

Questionnaire (84.7%)

173 Belgian and 10 Luxembourg participants

Level of clotting factors, %

1136393XII989354XI395425IX1188597VIII12197X212915VII7210576V233115II

888488838610

KULeuven, UCL Mont-Godinne

Survey 2008/3: PT reagents

216Total

2.82.8

661

Rabbit brain extractRecombinant rabbit TFRabbit brain extract

IL PT-Fibrinogen HSIL PT-Fibrinogen RecombinantIL PT-Fibrinogen

3.27Recombinant human TFIL Recombiplastin4.29Rabbit brain extractKordia Simplastin Excel S6.915Recombinant human TFRoche Neoplastin R7.917Human placentaSiemens Thromborel S13.930Recombinant human TFIL Recombiplastin 2G23.651Recombinant human TFSiemens Innovin34.374Rabbit brain extractRoche Neoplastin CI Plus%NThromboplastinReagent

Survey 2008/3: INR Results

2158.92.868.02.058.83.74Overall173.12.653.81.965.33.48Thromborel S84.93.063.22.112.53.91Simplastin Excel S745.32.933.92.125.13.90Neoplastin CI Plus 303.62.684.61.953.83.50Recombiplastin 2G76.03.104.02.247.83.89Recombiplastin154.93.244.02.246.54.06Neoplastin R515.62.705.21.916.13.51InnovinNCV,%INRCV,%INRCV,%INRReagent

CO/8884CO/8883CO/8610

Survey 2008/3: INR Results

3.193.062.202.114.263.91Simplastin Excel S

2.782.652.041.963.673.48Thromborel S

2.922.681.951.953.903.50Recombiplastin 2G

2.792.701.951.913.663.51Innovin

RELAC8884

INR 8884

RELAC8883

INR8883

RELAC8610

INR8610

Survey 2008/3: Questionnaire

• Citrate concentration• Lot number thromboplastin• Calculation vs calibration of INR• If calculation:

ISI: Manufacturer/Local calibrationNormal PT: MNPT/Manufacturer/

Pool/Lyophilized plasma…• Validation of INR

Citrate concentration for specimen collection

1.120.109/0.1293.870.129

67.01220.1098.8160.10619.2350.105

%Number of participantsCitrate mol/L

0.129 mol/L citrate

March 2000: 84.8%March 2005: 32.5%November 2006: 12.3%November 2008: 4.9%

Survey 2008/3: Questionnaire

• Citrate concentration• Lot number thromboplastin• Calculation vs calibration of INR• If calculation:

ISI: Manufacturer/Local calibrationNormal PT: MNPT/Manufacturer/

Pool/Lyophilized plasma…• Validation of INR

Lot number

• Different lot numbers in use

• For Recombiplastin 2G and Neoplastin CI Plus: enough participants to compare resultsobtained by 2 different lot numbers

Lot number

222.82.94194.32.928884

222.72.12193.32.148883

223.63.91192.43.888610Lot number 100905Lot number 100719Neoplastin CI Plus

123.02.68133.42.648884

122.71.98132.31.938883

123.43.49134.33.478610Lot number 573983Lot number 1172514Recombiplastin 2G

NCV,%INRNCV,%INR

Survey 2008/3: Questionnaire

• Citrate concentration• Lotnumber thromboplastin• Calculation vs calibration of INR• If calculation:

ISI: Manufacturer/Local calibrationNormal PT: MNPT/Manufacturer/

Pool/Lyophilized plasma…• Validation of INR

Calculation vs calibration (n=183)

Calculation of INR: 73.8%Calibration of INR: 26.2%

PT-Multi Calibrator (Dade-Behring)16/17 users (94.1%) of Thromborel S reagent30/41 users (73.2%) of Innovin reagentAccuclot (Kordia)1 user of Simplastin Excel S (Thrombolyzer)AK Calibrant (Technoclone)1 user of Neoplastin CI Plus (AMAX 200)

InnovinCalculation (11) vscalibration (30)

1,5

1,75

2

2,25

2,5

2,75

3

3,25

3,5

3,75

4

INR

Inn

ovin

PT-

Mu

lti C

al 8

610

Inn

ovin

861

0

Inn

ovin

PT-

Mu

lti C

al 8

883

Inn

ovin

888

3

Inn

ovin

PT-

Mu

lti C

al 8

884

Inn

ovin

888

4

3.47

1.90

2.70

3.72

2.0

2.86

Median CV,%

5.3 3.7

3.9 2.6

4.6 3.6

PT-Multi CalibratorInnovin (30) vsThromborel S (16)

1,5

1,75

2

2,25

2,5

2,75

3

3,25

3,5

3,75

4

INR

Inn

ovin

PT-

Mu

lti C

al 8

610

Thro

mbo

rel P

T-M

ult

i Cal

861

0

Inn

ovin

PT-

Mu

lti C

al 8

883

Thro

mbo

rel P

T-M

ult

i Cal

888

3

Inn

ovin

PT-

Mu

lti C

al 8

884

Thro

mbo

rel P

T-M

ult

i Cal

888

4

Median CV, %

3.47 3.47

1.90 1.95

2.702.655.3 5.9

3.9 3.8

4.6 3.1

Survey 2008/3: Questionnaire

• Citrate concentration• Lotnumber thromboplastin• Calculation vs calibration of INR• If calculation:

ISI: Manufacturer/Local calibrationNormal PT: MNPT/Manufacturer/

Pool/Lyophilized plasma…• Validation of INR

ISI

0

5

10

15

20

25

30

35

40N

umbe

r of

par

tici

pant

s

,8 ,9 1 1,1 1,2 1,3 1,4ISI value

ISI (n=133)

• Recommended by manufacturer: 92.5%

• Local ISI calibration: 7.5%

Local ISI determination (n=10)

1Patient samplesSTAR Recombiplastin 2G1Patient samplesSTAR EvolutionRecombiplastin1Patient samplesACL TOPPT-Fibrinogen HS1EtaloquickSysmex CA-540Neoplastin CI Plus1Patient samples Innovin/BCSACL TOPInnovin

1Patient samples Recombiplastin 2G/ACL TOP

ACL TOPInnovin1Patient samples ThrombotestBCS XPInnovin1Patient samples ThrombotestSysmex CA-1500Innovin2RELAC Sysmex CA-1500InnovinNCalibration plasmasInstrumentReagent

Innovin: ISI manufacturer vs local ISI determination

63.72.8654.82.80888464.22.0052.22.00888361.63.7354.73.638610NCV, %INRNCV, %INR

Local ISI determinationISI manufacturer

Homogeneous↔non-homogeneous (12)

1.8 2.0 2.2 2.4 2.6 2.8

3.0

3.5

4.0

4.5

5.0

Recombiplastin 2006/3

7395

7400

Homogeneous↔non-homogeneous (7)

1.8 2.0 2.2 2.4 2.6 2.8

3.0

3.5

4.0

4.5

5.0

Recombiplastin 2008/3

8883

8610

Homogeneous↔non-homogeneous(15)

2.0 2.2 2.4 2.6 2.8

3.0

3.5

4.0

4.5

5.0

Neoplastin CI PLUS 2006/3

7395

7400

Homogeneous↔non-homogeneous (4)

2.0 2.2 2.4 2.6 2.8

3.0

3.5

4.0

4.5

5.0

Neoplastin CI PLUS 2008/3

8883

8610

Homogeneous↔non-homogeneous (9)

1.8 2.0 2.2 2.4 2.6

2.5

3.0

3.5

4.0

4.5

5.0

Innovin 2006/3

7395

7400

Homogeneous↔non-homogeneous (3)

1.8 2.0 2.2 2.4 2.6

2.5

3.0

3.5

4.0

4.5

5.0

Innovin 2008/3

8883

8610

Survey 2008/3: Questionnaire

• Citrate concentration• Lotnumber thromboplastin• Calculation vs calibration of INR• If calculation:

ISI: Manufacturer/Local calibrationNormal PT: MNPT/Manufacturer/

Pool/Lyophilized plasma…• Validation of INR

Normal PT (n=131)

46.6% MNPT23.7% 100% value of reference curve19.8% Manufacturer

4.6% Pool3.1% Lyophilized plasma2.3% Combination

MNPT: Number of healthy individuals

1Infinite1Hundreds2500135022002101-15010100270450-59840-49330-39

2420-29N laboratoriesN healthy individuals

Normal PT

1/1/1/1MNPT/Pool/Lyophilized plasma/Combination

450.0 100% reference curveSimplastin Excel S1/1/1Pool/Manufacturer/Combination666.7MNPTNeoplastin R

2/1Pool/Combination261.712.946.4MNPT270.612.848.2100% reference curveNeoplastin CI Plus119.211.337.9MNPT188.110.862.1ManufacturerRecombiplastin 2G2Manufacturer571.4MNPTRecombiplastin

2/1Lyophilized plasma/Pool872.7MNPT InnovinNCV,%Median%PT normal plasmaReagent

Normal PT

264.72.933.12.135.83.92MNPT

274.42.944.02.113.53.88100% reference curveNeoplastin CI Plus

116.22.674.81.937.33.45MNPT

183.32.684.11.953.03.52Manufacturer

Recombiplastin 2G

NCV%

INR 8884

CV%

INR8883

CV%

INR8610

Reagent

Survey 2008/3: Questionnaire

• Citrate concentration• Lotnumber thromboplastin• Calculation vs calibration of INR• If calculation:

ISI: Manufacturer/Local calibrationNormal PT: MNPT/Manufacturer/

Pool/Lyophilized plasma…• Validation of INR

CLSI (H54)

• Validation of current INR testing should beperformed in each laboratory

• Minimum of 3 certified plasmas with a range of INRs between 1.5 and 4.5

• Validation should take place with any change in reagent, reagent lot number, instrument or following a major instrument repair

• INR obtained using the local system shouldcompare to the assigned INR value of the certified plasmas by +/- 15%

Validation of INR (n=172)

47 laboratoires (27.3%) mentionned validatingthe accuracy of their INR results

However: the majority of those laboratories refer toa validation of the accuracy of theirINR results by means of IQC

4 of them refer to participation in EQAS

ConclusionsInsights into the current practices in PT testing in Belgium and Luxembourg

All laboratories use low-ISI thromboplastins

High diversity of techniquesDifferent techniques yield different resultsWithin a given technique: small CVNo technique is significantly better than another

Compared to 2006/3: improvement of overall CVindicating a closer agreement between the INR results of the different laboratories

Conclusions: INR results

2158.83.748.92.868.02.05

NCV,%INR 8610

CV,%INR8884

CV,%INR8883

22412.73.8515.73.079.72.29

NCV,%INR 7400

CV,%INR7399

CV,%INR7395

Conclusions: Methodologies

• 95.1 % of the laboratories use 0.105-0.109 mol/L trisodium citrate as anticoagulant

• 93.1% of the laboratories use homogeneous test systems (reagent/instrument) ↔ 83.6% in 2006

• 46.6% determine the system-specific MNPT ↔37.3% in 2006

Validation of INR: Need for guidelines and control materials

Thanks to

EQA advisory boardJ. ArnoutB. ChatelainK. DevreeseK. JochmansO. PradierW. Wijns

AMHP van den BesselaarParticipating laboratories