Quality Controls in Allergy Diagnosis

Transcription

1 Quality Controls in Allergy Diagnosis Alistair Crockard Royal Hospitals Belfast Northern Ireland

2 Quality Controls in Allergy What do we want? Diagnosis What can be controlled? What can be achieved?

3 What do we want? 1. A process that ensures that: the results generated are correct

4 What do we want? 1. A process that ensures that: the results generated are correct QUALITY CONTROL

5 What do we want? 2. A process that ensures: the quality of results is consistent and in line with those determined for recognised reference material (standards)

6 What do we want? 2. A process that ensures: the quality of results is consistent and in line with those determined for recognised reference material (standards) QUALITY ASSESSMENT

7 What do we want? 3. A process that ensures: the correc test is carried out on the correct sample and the correct result, with the correct interpretation, is delivered to the correct person at the correc time.

8 What do we want? 3. A process that ensures: the correc test is carried out on the correct sample and the correct result, with the correct interpretation, is delivered to the correct person at the correc time. QUALITY ASSURANCE

9 What do we want? 4. Guidelines: CLSI Approved Guidelines I/LA20-4 (1997) Evaluation methods and analytical performance characteristics of immunology assays for human IgE antibodies of defined specificities 2 nd Edition due late

10 What can be controlled? Steps in the Analytical Process

11

12 What can be controlled? Steps in the Analytical Process Blood Sample Assay Results Report correct patient correct sample correct request instrument reagents method assay performance validation of results interpretation

13 What can be controlled? outside and inside the laboratory Outside Instrumentation / Assay Assay Methodology Detection System Assay Sensitivity Reagents Preparation Purity Allergenicity Reports Clinical Interpretation

14 What can be controlled? outside and inside the laboratory Outside Instrumentation / Assay Assay Methodology Detection System Assay Sensitivity Reagents Preparation Purity Allergenicity Reports Clinical Interpretation Inside Assay Performance Precision Accuracy Reproducibility Quality Control Quality Assessment Result Interpretation

15 What can be achieved? Outside the Laboratory Instrumentation / Assay Methodology / Reagents Commercial Companies Extensive Quality Programmes

16 Quality Control procedures for components used in Specific IgE assay Serum Allergen Raw material Raw Sponge Selected control sera Antibody Activated Sponge Enzyme ImmunoCAP IgE Calibrator WHO reference preparation Conjugate Buffer Software Instrument Assay Development Solution aige ImmunoCAP Antibody Activated Sponge Wash Solution Stop Solution Buffer Substrate Raw Sponge Quality Control / methods and specifications

17 What can be achieved? Outside the Laboratory Reagents Heterogeneous starting material Variable extraction/purification/synthetic procedures Natural / recombinant allergens Material of variable allergenicity Standardisation problematic

18 What can be achieved? Outside the Laboratory Allergen Standardisation limited WHO International Reference Preparation 75/502 for human IgE generally adopted as industry standard WHO International Reference Preparations for birch pollen; dog hair/dander; HDM (D pteronyssinus); ragweed pollen; timothy grass pollen generally ignored as industry standard Units: kiu/l and ku A /l not equivalent

19 d1 e1 g6 t3

20

21 CREATE Project Consortium of interest groups: Clinical; Research; Industrial; Regulatory Task: Produce purified recombinant and natural allergens Evaluate recombinant major allergens as candidate certified reference materials Validate sandwich ELISAs for quantitation

22

23 Panel of 9 recombinant molecules studied 3 molecules identified which displayed good structural and immunological reactivity and biological potency: rbet v1; rphl p 5a; rder p 2 ELISAs verified rbet v 1 and rphl p 5a established as international reference standards by end 2008

24 What can be achieved? Inside the Laboratory Monitoring Assay Performance Implementing effective internal QC schemes Incorporating control sera, of defined specific IgE concentrations, into each assay run Plotting performance using Shewhart / Levy- Jennings plots

25 Shewhart Chart 100 KU/l sd +2 sd +1 sd Target value -1 sd -2 sd -3 sd Assay Run Allergen D1: Target Value = 49 ku/l

26 Step Change Bias Drift In Control

27 What can be achieved? Inside the Laboratory Monitoring Assay Performance Implementing effective internal QC schemes Incorporating control sera, of defined specific IgE concentrations, into each assay run Plotting performance using Shewhart / Levy- Jennings plots Applying Westgard rules to determine assay acceptability

28 Westgard Rules Used to define define specific performance limits for an assay Detect both random and systematic errors 6 commonly used rules 3 warning rules (1 2SD ; 2 2SD ; 4 1SD ) 3 mandatory (1 3SD ; R 4SD ; 10x)

29 Westgard Rules Violation of warning rules should trigger a review of test procedures, reagent performance and equipment calibration Violation of mandatory rules should result in the rejection of the results obtained with patients serum samples in that assay

30 Monthly IQC Peanut (F13) March 2008 Value ASSAY: peanut f13 (Immulite DPC) Date kit kit expiry allergen allergen expiry Control A Control B Rule Violations CONTROL DATA COMMENTS lot no date lot no date Value SD Value SD 1 2S 1 3S 2 2S R 4S 4 1S 10 X and carry over Control A Accept (GB) Batch Accept (GB) Mean SD CV% Accept (GB) A Accept (GB) Mean - Accept (GB) Accept (GB) Last 3 SD's Accept (GB) th Accept (GB) th Accept (GB) th Accept (GB) Control B Accept (GB) Batch Accept (GB) Mean SD CV% Accept (GB) Accept (GB) Mean - Accept (GB) Accept (GB) Last 3 SD's Accept (GB)

31 What can be achieved? Inside the Laboratory Particular difficulties in running allergen specific IgE IC schemes Limited availability of reference sera and commercial preparations Limited availability of in house serum samples of clinically relevant concentrations: CVs concentration dependent Resulting in the inability to run IQ for all allergenspecfic IgE assays Use data from representative allergen-specfic IgE assays

32 What can be achieved? Inside the Laboratory Monitoring Assay Performance Participating in external Quality Assessment schemes Comparative information about methods and analytical platforms Providing a measure of performance Commercial companies Distribution of sera of defined concentrations Internet-based service and support Independent providers Distribution of sera of defined concentrations Case histories / interpretative reporting

33 What can be achieved? Inside the Laboratory Monitoring Assay Performance Participating in external Quality Assessment schemes Comparative information about methods and analytical platforms Providing a measure of performance Commercial companies Distribution of sera of defined concentrations Internet-based service and support Independent providers Distribution of sera of defined concentrations Case histories / interpretative reporting

34

35 What can be achieved? Inside the Laboratory Monitoring Assay Performance Participating in external Quality Assessment schemes Comparative information about methods and analytical platforms Providing a measure of performance Commercial companies Distribution of sera of defined concentrations Internet-based service, support and QC programmes Phadia LabCommunity ; Siemens Real Time Solutions Independent providers Distribution of sera of defined concentrations Case histories / interpretative reporting

36

37

38

39 What can be achieved? Inside the Laboratory Monitoring Assay Performance Participating in external Quality Assessment schemes Comparative information about methods and analytical platforms Providing a measure of performance Commercial companies Distribution of sera of defined concentrations Internet-based service, support and QC programmes Phadia LabCommunity ; Siemens Real Time Solutions Independent providers Distribution of sera of defined concentrations Case histories / interpretative reporting

40 Single donor samples with case histories Medical history Physical examination Physiological investigations Allergy investigations (SPT; challenge tests) Conclusion / Diagnosis

41 Sample Allergic rhinitis IgE sensitisation to HDM Grass pollen Animal dander Latex IgE

42 Sample Allergic rhinitis IgE sensitisation to HDM Grass pollen Animal dander Latex e1

43 Sample Allergic rhinitis IgE sensitisation to HDM Grass pollen Animal dander Latex e3

44 Sample Allergic rhinitis IgE sensitisation to HDM Grass pollen Animal dander Latex k82

45 Schemes IgE Allergen-specific IgE; 15 common allergens Single donor samples Performance assessment of quantitative responses and consensus of results by grades

46

47

48 Web-based educational tool Allows individuals to practice clinical and scientific interpretation skills on virtual patient results Navigate through screens of test results and medical information Suggest tests and investigations; draw conclusions Compare with correct answers Award certificate of participation

49

50

51

52

53

54 Cellular Tests

55 Cellular Tests Basophil Activation Allergen CD63+ CD203c+ Activated Basophil Anti-IgE

56 Basophil Activation Tests Standardisation Protocols Technical Aspects Interpretation

57 EuroBAT Project Initiative to establish the diagnostic validity of BAT in clinical scenarios where current tests are: Unreliable Difficult to perform Unavailable

58 EuroBAT Project Undertake multicentre studies to validate Protocols Reagents Commercial kits Produce position papers / guidelines Develop European standard reference protocols Maintain a website

59 EuroBAT Working Groups Technical Issues Venom Food Autoimmune Urticaria Drugs Betalactams NSAIDS Muscle Relaxants

60 Quality Controls in Allergy What do we want? Diagnosis What can be controlled? What can be achieved?

61 Quality Controls in Allergy What do we want? Diagnosis What can be controlled? What can be achieved? What more to do?

62 Future Challenges Cellular Assays Micro Array Assays

63 Future Challenges Recombinant Allergens EMBO 2004

64 Future Challenges Recombinant Allergens Recombinant Antibodies I. Braren et al Clin Chem : 837 EMBO 2004

65 Future Challenges Near Patient Testing

66 Patient Responses Genetic Diversity

67 Patient Responses Genetic Diversity 50 (something) year old atopic male Occasional rhinitis Specific IgE HDM: Positive 6 ku/l SPT HDM: Positive 4mm wheal Basotest HDM: Positive ImmunoCAP TM Rapid: NEGATIVE

68

69 Thank you for your attention