Le point sur les techniques de g notypage plaquettaire le bilan du registre suisse hpa hla
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Le point sur les techniques de génotypage plaquettaire & le bilan du registre suisse HPA / HLA. Françoise Boehlen Médecin adjointe, CC Service d ’ angiologie et d ’ hémostase HUG. Swisstransfusion – 6 septembre 2013. Presentation. Human platelet antigens and tests

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Le point sur les techniques de génotypage plaquettaire & le bilan du registre suisse HPA / HLA

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Le point sur les techniques de g notypage plaquettaire le bilan du registre suisse hpa hla

Le point sur les techniques de génotypage plaquettaire & le bilan du registre suisse HPA / HLA

Françoise Boehlen

Médecin adjointe, CCService d’angiologie et d’hémostase

HUG

Swisstransfusion – 6 septembre 2013


Presentation

Presentation

  • Human platelet antigens and tests

  • Clinical aspects of platelet alloimmunisation

  • National HPA / HLA platelet registry


Platelet alloantigens

Platelet alloantigens

Two different categories of clinically relevant platelet alloantigens:

  • « Common alloantigens » found on platelets and other blood cells or tissues (e.g. antigens of the ABO-system and HLA-class I antigens)

  • « Platelet-specific antigens » = HPA (Human Platelet Antigen), thought to be present exclusively on platelets


Le point sur les techniques de g notypage plaquettaire le bilan du registre suisse hpa hla

GPIV

GPVI

CD9

GPIIb-IIIa

a6b1

a5b1

GPIa-IIa

GPIb-IX-V

Platelet membrane


Gpiib iiia

GPIIb-IIIa

Fibrinogenbinding site

GPIIIa

GPIIb

RGDbinding site

Ca++

Ca++

Ca++

HPA-4

Ca++

HPA-1

HPA-3

s

s

HPA-1aa

HPA-1ab

HPA-1bb

COOH

COOH


Tests

Tests

  • HPA typing

  • Antiplatelet antibodies detection

  • External Quality Controls (tests performed in HUG)

  • NIBSC (National Institute for Biological Standards and Control)

  • International Platelet Immunology Workshop (ISBT)


Platelet typing

Platelet typing

  • Phenotyping methods (serologic typing)

    • Different methods (immunofluorescence, MAIPA…)

    • Relatively easy to perform but depend on the availability of appropriate human sera

  • Genotyping methods

    • Extraction of genomic DNA from blood leukocytes (or amniocytes)

    • Amplification by PCR (different techniques: RFLP, SSP, etc.)


Example

Example

1a

1b

2a

2b

3a

3b

5a

5b

+

-

+

-

+

-

+

+

Platelet genotype

1aa 2aa 3aa 5ab


Antiplatelet antibodies detection

Antiplatelet antibodies detection

  • MAIPA (monoclonal antibody-specific immobilisation of platelet antigens)

    • Gold standard reference technique in platelet immunology

  • Identification of specific platelet antibodies

    • GPIIb-IIIa, GPIb-IX, GPIa-IIa, etc. (HLA class I)

  • Characterisation of platelet alloantibodies by cross-match between

    • Mother’s serum and father’s platelets

    • Patient serum and panel of platelets of different genotypes


Presentation1

Presentation

  • Human platelet antigens and tests

  • Clinical aspects of platelet alloimmunisation

  • National HPA / HLA platelet registry


Alloimmune thrombocytopenia

Alloimmune thrombocytopenia

Platelet alloantibodies are responsible for:

  • NAITNeonatal alloimmune thrombocytopenia

  • PTPPosttransfusion purpura

  • PAITPassive alloimmune thrombocytopenia

  • TAITTransplantation-associated alloimmune thrombocytopenia

    Platelet alloantibodies may contribute to:

  • PTRPlatelet transfusion refractoriness


Alloantibodies

Alloantibodies


Presentation2

Presentation

  • Human platelet antigens and tests

  • Clinical aspects of platelet alloimmunisation

  • National HPA / HLA platelet registry


Hpa registry of blood donors

HPA registry of blood donors

  • Swiss registry of HLA- and HPA-genotyped platelet donors

    • Proposed by Swiss Transplant Working Group for Blood and Marrow Transplantation (STABMT)

    • Goal = 2’000 platelet donors HPA genotyped

    • Financed by

      • Swiss Red Cross Humanitarian Foundation

      • Blood Transfusion Service, Swiss Red Cross


Project part 1

Project – Part 1

Platelet Apheresis Centres

Selection of regular apheresis and blood donors already HLA-typed and < 50 years old

Informed consent sheet  questionnaire (donors data)

Blood sample


Project part 2

Project – Part 2

Laboratory, Haemostasis Unit, Geneva

HPA-genotyping

Anti-HPA ab screening in homozygous women with ≥ one pregnancy or 2nd/3rd trimester miscarriage

Introduction of results in the registry (website) after quality control


Project part 3

Project – Part 3

Platelet Apheresis Centres

Documentation of HPA-matched transfusions

Update of the list of donors (contraindication)

Inclusion of new donors


Website

Website

www.hpa-hla.org

Only for participating centres


Registry

Registry


Example mrs y 50 year old

Example – Mrs Y, 50 year-old

  • Acute myeloid leukaemia

  • Platelet transfusion refractoriness with bleeding

  • Anti-HLA class I & II, and anti-HPA-5a alloantibodies

    • Patient platelet genotyping: HPA-5bb

    • Persisting refractoriness including after HLA-matched HPA-5-mismatched products

  • Myeloablative allogenic HSCT from her HLA-identical brother

    • Brother platelet genotyping: HPA-5ab

    • Selection HLA- and HPA-5 compatible platelet donors


Example registry

Example – Registry


Registry statistics

Registry – Statistics

  • Beginning of the project in March2003

     3113 samples received at the end of August 2013

31

717

2116

249


Centres 2013

Centres (2013)


Registry annual statistics

Registry – Annual statistics

Last 12 months (from 01.08.2012 to 31.07.2013)

  • No documentation of HPA-matched transfusions

    • How many platelet donors searched/found?

    • How many products prepared/transfused?

    • For which reason?


Rare platelet genotypes

Rare platelet genotypes

 2116 donors available at the end of August 2013

2.5%

0.8%


Registry characteristics

Registry – Characteristics


Summary

Summary

  • Aim of this registry

    • HPA-compatible platelet in case of

      • Neonatal alloimmune thrombocytopenia

      • Platelet transfusion refractoriness

      • (Posttransfusion purpura)

  • Many questions to be resolved

    • Real necessity?

    • Necessity of a huge registry to propose HLA- and HPA-compatible platelets?

    • What about detection of alloantibodies in blood donors?


Thanks

Thanks

Prof. Philippe de Moerloose

Head of Haemostasis Unit

Dre Cécile Kaplan & coworkers

INTS, Paris

Oana Bulla & Yen Lai

Technician

Claude-Alain Mouthon

Website designer

Drs Rudolf Schwabe & Guy Levy

Drs Grazia Nicoloso de Faveri & Behrouz Mansouri

Blood Transfusion Service SRC

Transfusion centres & donors


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