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1. Background
Platelet immunology continues to develop at a rapid pace and the number of clinically relevant platelet specific alloantigen systems (HPA) has now increased to seventeen. Platelet specific alloantibodies are involved in neonatal alloimmune thrombocytopenia (NAITP), platelet refractoriness and post-transfusion purpura (PTP).
The aim of the Platelet Immunology Quality Exercises is to help those clinical laboratories concerned with the detection of platelet-specific antibodies achieve a high standard of performance, by distributing coded samples for testing.
2. Platelet Immunology Exercises
The scheme has two parts, a) the detection and identification of platelet alloantibodies and b) the detection of HPA alleles by molecular techniques. Laboratories may opt for either scheme or both. Currently there are 34 participants in the serology scheme and 24 in the HPA genotyping scheme. The proportion of non-UK participants has gradually increased, as shown below;
Serology Exercise
UK
Europe
Australia
North America
Total
1998-A
14
13
3
1
31
2005-A
9
20
2
34
Genotyping Exercise
10
0
12
6
16
28
3. Organisation
There are two exercises each year, organised by a Steering Committee under the umbrella of NIBSC. In recognition of the wider participation by laboratories outside the UK the Steering Committee is currently inviting members from other countries to join the Steering Committee. The current membership of the committee is;
Dr Willem Ouwehand NIBSC & University of Cambridge (chairman)
Mr Dave Allen National Blood Service, Oxford
Dr Riitta Kekomaki Red Cross, Finland
Dr Cecile Kaplan INTS, France
Dr Masja De Haas CLB, The Netherlands
Dr Paul Metcalfe NIBSC, Potters Bar
Members are appointed for three years and can be re-elected for one additional term. Replacement is on a rolling basis. The committee meets twice yearly to discuss current performance and plan future exercises. Applications to join the scheme are considered by the committee and continued participation depends on timely submission of results.
4. Samples distributed
Serology
Usually four 2.0ml coded samples are distributed. Samples included in the exercises are generally related to genuine clinical cases. It is not standard policy to include mixtures of different samples, although dilution of samples with potent antibodies is sometimes required to provide enough material for evaluation. All samples are evaluated at NIBSC before distribution. Selected samples are aliquoted and distributed along with instructions for testing and results sheets. Samples to UK Participants are despatched at ambient temperature by First Class post, samples to other european laboratories are delivered at ambient temperature by 'door to door' courier and samples to non-european labs are delivered on dry-ice by 'door to door' courier. Participants are usually asked to test four coded samples using their routine in-house techniques for the detection of platelet alloantibodies.
Genotyping
Four 1-1.2ml EDTA anti-coagulated whole blood samples are distributed at the same time as the serology samples. Laboratories are expected to determine the HPA- 1, -2, -3 -5 & -15 genotypes of the four samples.
5. Assessment
There is considerable inter-laboratory and inter-exercise variation in alloantibody detection 1. Performance in HPA genotyping has improved considerably and is now generally excellent; some laboratories report single errors, but most are entirely correct 1.
Results must be returned within two weeks, a sample de-code is issued one week after the closing date and a report detailing the results of all laboratories (anonymous) is distributed at a later date. All laboratories are tested on their ability to detect and identify HPA antibodies. HLA antibodies may be present in the samples but their detection is not included in the scoring scheme.
Scoring system
An antibody must be detected by NIBSC and at least four other laboratories to be included in the scoring scheme. The scoring scheme has been adapted from other serology proficiency schemes in the UK; points are awarded for correct determination of the presence or absence of an antibody and points are subtracted for an incorrect answer. A percentage mark is then calculated from the maximum marks available. A mean score over the last three exercises is calculated for each laboratory. HPA genotyping is scored in a similar way.
After each exercise a report is distributed to all participants. Written feedback from participants to the Steering Committee is encouraged and many of the changes to the format of the exercises have come in response to suggestions from participants. An official response will be returned to the participant after the Steering Committee has convened.
The results of the exercises are published at regular intervals in peer-reviewed journals1 and consensus protocols are placed on the NIBSC Website.
Poor performance
Poor performers (the cut off for poor performers is 75% in the serology scheme and 95% for HPA genotyping) are contacted by the Scheme Organiser with the aim to try to identify ways which could lead to improved performance.
6. Cost
To cover the cost of materials and distribution of the serology exercises a charge is made to clinical laboratories of £137 per exercise (+ vat if applicable) for the serology section, and £85 per exercise (+ vat if applicable). Commercial companies are charged double.
7. Associated activities
Occasionally other materials for testing are distributed with the serology exercise e.g. the WHO Reference Reagent anti-HPA-5b freeze dried preparation for sensitivity determination, or other candidate materials for reagents or standards, in which case the number of samples for 'blind testing' is reduced.
On other occasions methodology questionnaires have been distributed so that details of laboratory procedures and reagents can be summarised and made available to participants. In this way consensus methods for antibody detection by PIFT and MAIPA and HPA genotyping by PCR-SSP have been established and made available.
8. Reference
1. D Allen, WH. Ouwehand, M de Haas, R Kekomaki, C Kaplan, P Metcalfe. Interlaboratory variation in the detection of HPA-specific alloantibodies and in molecular HPA typing. Vox Sanguinis 2007, 93, 316-324.