Building a haemoglobinopathy library
Haemoglobinopathies are a diverse and heterogeneous group of disorders that affect the genes responsible for synthesising globin, an essential component of haemoglobin. They are considered to be one of the most common autosomal recessive disorders affecting humans and can affect either the type of globin produced (haemoglobin variants) or the amount (thalassaemia). In adults, the majority of globin is synthesized by 2 genes, the beta globin gene and the alpha globin gene, both are essential because the main adult haemoglobin (Hb A) is comprised of tetramers formed of two alpha and two beta chains. Although there are a number of common haemoglobin variants such as Hb S, Hb C and Hb E, over 1,000 haemoglobin mutations have been reported.
Most laboratories can identify the common variants but identification of the rare variants is difficult without DNA analysis.
Several laboratory tests are available to screen for haemoglobin variants, such as high performance chromatography (HPLC), capillary electrophoresis (CE), alkaline and acid agarose gels, iso electric focusing (IEF), and others. However these methods will only be able to produce a presumptive identification.
With the advances of genetic testing, DNA analysis provided a reliable confirmatory method detecting mutations ranging from a single nucleotide alteration to the loss, duplication or rearrangement of several thousands of nucleotides. An example of these techniques is DNA sequencing by Sanger sequencing.
But, due to the price and processing time, these DNA based methods can’t be used routinely to screen all patients for haemoglobin variants and thalassaemias.
Building the resource
Companies try to help the laboratories to provisionally identify variants by offering online library facilities which identify the characteristics of different variants with the equipment in use. This provides a helping hand for the presumptive identification of haemoglobin variants and what further testing is required.
Viapath laboratories offers molecular techniques for all globin genes and mass spectrometry to identify haemoglobin variants, alongside the relevant expertise required for the interpretation of this complex area.
Trinity Biotech (Bray, Co. Wicklow, Ireland) selected Viapath’s Special Haematology Department at Guy´s and St. Thomas NHS Foundation Trust for a joint project, aimed at building an online haemoglobinpathy library resource for their Premier Resolution analyser (HPLC). All haemoglobinopathies characterised in the library are confirmed by DNA or mass spectrometry analysis much of which has been carried out at the Viapath laboratories. Furthermore with this partnership, Trinity Biotech can provide a haemoglobinopathy confirmation service for their clients all over the world.
There are a large number of haemoglobinopathies and their combinations. Therefore building the library requires a large number of samples which have been gathered from several countries. Trinity Biotech is shipping samples, they have collected and preserved over the years, to Viapath’s laboratory where the necessary testing can be performed to confirm the genotype.
After confirmation, the results are sent back to Trinity Biotech where they are integrated into the library and facilitated to the end user – the operators of their Premier Resolution Analyser. When an abnormality is found within a given sample, the software compares the peak characteristics present in the patient sample against the examples in the library, the software then suggests a number of possibilities in order to aid the operator in the result interpretation.
The library is comprised of chromatography plots available for each of the samples confirmed by DNA analysis, but also provides information about: the name and characteristics of each Haemoglobinopathy, its retention time (RT), the globin chain affected and the family origins where they can be found. An example is shown in the schematic below.
For further information, please contact:
Yvonne Daniel, Lead Scientist and Operational Lead, Haematology
Yvonne [dot] Daniel [at] viapath [dot] co [dot] uk
Daniel Monteiro, Senior Biomedical Scientist
Daniel [dot] Monteiro [at] viapath [dot] co [dot] uk