Relationship between neonatal screening results by HPLC and the number of α-thalassaemia gene mutations; consequences for the cut-off value.

Objectives: To evaluate the relationship between FAST peak percentage by adapted Bio-Rad Vnbs analysis using the valley-to-valley integration and genotypes with the aim to improve differentiation between severe α-thalassaemia forms (HbH disease) and the milder disease types.
Method: DNA analysis for α-thalassaemia was performed on 91 dried blood spot samples presenting normal and elevated FAST peak levels, selected during three years of Dutch national newborn screening.
Results: Significant differences were found between samples with and without α-thalassaemia mutations, regardless of the genetic profiles. No significant difference was demonstrated between HPLC in -α/αα and -α/-α, between -α/-α and - -/αα and between - -/αα and - -/-α genotypes.
Conclusion: This study confirms that the percentage HbBart's, as depicted by the FAST peak, is only a relative indication for the number of α genes affected in α-thalassaemia. Based on the data obtained using the modified Bio-Rad Vnbs software, we adopted a cut-off value of 22.5% to discriminate between possible severe α-thalassaemia or HbH disease and other α-thalassaemia phenotypes. Retrospectively, if this cut-off value was utilized during this initial three-year period of neonatal screening, the positive predictive value would have been 0.030 instead of 0.014.