quality in non- infectious serology · 2015. 6. 11. · martine vercammen national symposium on...

Martine Vercammen National Symposium on Non-Infectious Serology 2015

Martine Vercammen

Quality in non-infectious serology


Martine Vercammen

Willy Vandersteen


Martine Vercammen

o Method » Validation/Verification o Training of personnel

o Monitoring o Short term o Long term

o Pre-analysis o Method o Pipetting and incubation

station o Reagents o Microscopy/Signal detection o People o Informatics o Post-analysis o Murphy?


Martine Vercammen

• Choice of method • Reagent Lot change • QC

• First level - IQC • Second level • Third level - EQC

• Key Performance Indicators


Martine Vercammen

National Symposium on Non-Infectious Serology 2015

Choise of a new method


Martine Vercammen

Performance Practical considerations

• EQC

• Literature

• Colleages

• Personnel and organisation

• Available equipment

• Reagent stability

• Sample number

• Frequency of analysis

• Turn-around time

• Finances


Martine Vercammen

» 5 most important patterns: 5 sera/pattern (known diagnosis) » Titration » 10 negative sera » Reproducibility

homogeneous speckled nucleolar centromere cytoplasmatic


Martine Vercammen

» 3 patterns: 5 sera/pattern (known diagnosis) » 10 negative sera » Reproducibility » Titration

C-ANCA P-ANCA Atypical ANCA


Martine Vercammen

Qualitative Quantitative

• min 5 sera/antigen

• 10 negatives

• Routine in duplo

• Reproducibility

• ≥ 10 patiënts/antigen

• Concentration range

• Routine in duplo

• (Evaluation cut-off)

• Reproducibility


Martine Vercammen

Lot change


Martine Vercammen

homogeneous

important patterns: 1 serum/pattern 1 negative serum 1 titration

speckled nucleolar centromere cytoplasmic

C-ANCA P-ANCA atypical ANCA


Martine Vercammen


Martine Vercammen

» 1/antigen or 1 pool of antigens » 1 negative serum


Martine Vercammen

IQC jump control material

comparison

patient samples

if clinically relevant

contact supplier


Martine Vercammen

Patient DWR Patient DWR


Martine Vercammen


Quality control


Control 200+/- 30

Patients high

Control 200+/-30


Martine Vercammen

1. Positive control - IF: 1 important pattern - immuno-assay: 1/antigen, alterning in case of multiplex - Dilution and handling: exactly equal to patient samples - Origin: patiënt or commercial, preferentially with known

pathology stability: 4°C 0.1 g/% Na azide/ frozen aliquots - Titration/Intensity score

2. Negative control if small series 3. Known patients: IF pattern/specificity/intensity


Martine Vercammen

• Each run: start and end

• Material: commercial or patient

• Acceptance criteria: company or laboratory


Martine Vercammen


Martine Vercammen


IQC Automatic microscope: role for the fluorescence intensity?


Martine Vercammen

» Reproducibility ˃ Within-run

o Speckled 1,4% G-sight Bonroy, Clin Chem Lab Med 2013

o Homogeneous 10,9% o Centromere 16% o Nucleolar 5% o SSA 15% o Scl-70 2 - 25% company-dependent Bizarro, JAutoimmunity Rev 2014

G-sight, NovaView, ImageNavigator, EuroPattern, Aklides Between run

o Homogeneous 16% G-sight Bonroy, Clin Chem Lab Med 2013

o Negative control 48%

˃ Between-labs? to be developed

» End-point titers


Martine Vercammen

Zenit G sight

Maenhout, Bonroy et al. Clin Chem Lab Med 2014; Bossuyt, Cooreman et al Acta Clinica Chimica Acta 2013; 2014


Martine Vercammen Schouwers Clin Chem Lab Med 2013

Nova view


Martine Vercammen


Peng, Tang et al. J Immunol Methods

EUROIMMUN reagents


Martine Vercammen

» Statistics on parameters that regulate sensitivity of the camera

» Time to focus related to 1/ fluorescence intensity



Martine Vercammen Maenhout, Bonroy et al. Clin Chem Lab Med 2014

CV= SD x 100 mean CV 2 % CV 41%

nucleolar

Westgard rules Warning 1S2 Rejection 2S2 1S3 R4S


Martine Vercammen Maenhout, Bonroy et al. Clin Chem Lab Med 2014


Martine Vercammen

Standardization between instruments o Intensity o Cut-off for positivity


Martine Vercammen

Personnel: tuning and evaluation of competence 1/year


Martine Vercammen

» = Proficiency testing » External QC

» Frequency » - minimum WIV/IPH programm (RIZIV/INAMI) » - preferentially 1 programm/test/year » (ISO 15189 accreditation)


Martine Vercammen


Martine Vercammen

Maenhout, Bonroy et al. Clin Chem Lab Med 2014

2S2

1S3

2S2 1S3


Martine Vercammen


Martine Vercammen

2008 2009 2010 2011 2012 2013 2014

Positive/total (%)

39 36 23 23 22

?


Martine Vercammen

Homogeneous Speckled Centromere Cytoplasmatic Nucleolar

?


Martine Vercammen

2010 2014

DNA ENA %

Expected Ag %

DNA ENA

%

Expected Ag %

homogeneous 21 7 dna histones 29 12

dna histones

speckled 16

13 SSA SSB Ro52 RNP SmD ribP

Scl70

36

25 SSA SSB Ro52

RNP SmD PM/Scl Scl70

centromere 85 79 Cenp B 100 100 cenp B

cytoplasmatic 8 1 Jo 1 17 10 Jo1 M2

nucleolar 7 4 RNP Scl70 1 0,25 Scl70

Different Hep2 cells Different ENA blot



Marc Sleen

quality in non- infectious serology · 2015. 6. 11. · martine vercammen national symposium on...

Documents