An international trial demonstrates suitability of a newly developed whole-blood ELISA kit for multicentre platelet HPA-1 phenotyping.
BACKGROUND
Neonatal alloimmune thrombocytopenia (NAIT) is in most cases due to pregnant women with HPA-1b platelet phenotype producing antibodies to HPA-1a platelets of the fetus, which may lead to intracranial haemorrhage with subsequent death or life-long morbidity. The availability of sensitive, reliable, straightforward, and inexpensive assays would enable large-scale screening in pregnancy and may help avoid NAIT.


METHODS
A recently developed enzyme-linked immunosorbent assay (ELISA) was produced in kit form incorporating modified reagents and enabling distribution to 21 international Platelet Immunology and Blood Centres (see Acknowledgements). The kits were assessed using anticoagulated whole-blood samples stored up to 10 weeks at 4 degrees C. Each centre tested its own blood samples most of which had been previously typed by established assays.


RESULTS
Of the 152 samples that were tested, all 31 HPA-1b phenotypes were correctly identified by the kit. The respective mean +/- standard deviation of the specific absorbances of HPA-1b and HPA-1a samples were: 0.52+/-0.15 and 0.12+/-0.06 (p<0.0005). In addition, the modified ELISA showed 100% concordance with PCR-SSP in the phenotyping of 93 donors. Finally, a comparison between freshly prepared and stored kits showed that the kit was stable for at least 2 years at 4 degrees C.


CONCLUSIONS
The international trial showed that the modified whole-blood ELISA kit is very well suited for wide-scale screening in pregnancy. Moreover, the ELISA kit could be used for large-scale phenotyping of blood donors, with HPA-1b-typed individuals getting invited to become apheresis platelet donors for patients with NAIT.
Bessos
H
H
Academic Transfusion Medicine Unit and South-East Scotland Blood Transfusion Service (BTS), Edinburgh, UK. Bessos@compuserve.com
Hofner
M
M
Salamat
A
A
Wilson
D
D
Urbaniak
S
S
Turner
M L
ML
D000328:Adult; D016824:Antigens, Human Platelet / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006801:Humans; D007231:Infant, Newborn; D039661:Integrin beta3; D007518:Isoantibodies / Q000276:immunology; D010641:Phenotype; D011247:Pregnancy; D011933:Reagent Kits, Diagnostic; D013921:Thrombocytopenia / Q000276:immunology
D016824:Antigens, Human Platelet; C061248:ITGB3 protein, human; D039661:Integrin beta3; D007518:Isoantibodies; D011933:Reagent Kits, Diagnostic; C083451:human platelet antigen 1b