Coexistence of hemoglobin Handsworth and alpha 3.7 kb deletion in Caucasian woman in Poland

BackgroundThis report presents a case of an adult Polish women of Caucasian origin who was heterozygous for the nondeletional mutation: Hb Handsworth (HBA2 or HBA1: c.55G > C, p.Gly19Arg) and deletional (-α3.7) α-thalassemia mutation.MethodsThe HbA2 and HbF levels were measured by microcolumn chromatography and alkaline denaturation procedure, respectively, while electrophoresis was used to detect pathological hemoglobin fraction. The β- and α-globin genotypes were determined by DNA sequencing, gap-polymerase chain reaction, α gene triplication and MLPA.ResultsThe HbA2 and HbF levels were normal, but hemoglobin electrophoresis on agarose gel alkaline pH showed a strong band migration in a position of hemoglobin S and faint bands in the neighborhood of band A on acid electrophoresis. Molecular analysis of the alpha globin cluster detected a point mutation at codon 19 in HBA2 (c.55G > C, p.Gl- y19Arg) and deletion -α3.7.ConclusionsOur compound heterozygosity does not produce severe clinical or hematological symptoms but it is important to say that in our part of Europe such cases do appear. Molecular analysis of the alpha globin cluster is required for correct diagnosis in patients with normal HbA2 levels. Compound heterozygosity was unmasked by molecular diagnosis only.

RAPID DETECTION OF -THALASSEMIA MUTATIONS IN THAILAND USING MULTIPLEX ARMS

The number of mutations underlining b-thalassemia generate a wide variety of different clinical phenotypes. An understanding of the genotype is important for medical personnel in order to provide proper counseling to patients and their families. Characterization of these mutations should aid the planning of a prenatal diagnosis program for bthalassemia. The heterogeneity of the mutations makes it difficult and time consuming to identify the mutation in some individuals. We developed a single-tube multiplex amplification refractory mutation system (multiplex ARMS) to identify common ethnic- specific b-thalassemia mutations. Confirmation of multiplex ARMS results was carried out using direct sequencing. Three thousand three hundred twenty two people from Phitsanulok province were screened for the b-thalassemia trait by quantitation of HbA2 with microcolumn chromatography and the genotypes of mutations were characterized using multiplex ARMS and direct sequencing. We found that the deletion at codons 41/42 (-TCTT) was the most frequent (48%), codon 17 (A®T) (30%), -28 (A®G) (6%) and IVS-I-1(G®T) (6%) were the second and third in frequency respectively. A -87 (C®A) mutation (4%), IVS II-654 (C®T) (2%), codons 71/72 (+A) (2%) and codon 35 (C®A) mutations (2%) were also found. These techniques were found to be a valuable tool for analysis of b-thalassemia mutations because they are accurate, simple, and speedy in operation. The application for the diagnosis of severe thalassemia in high-risk pregnancies is promising.

Capillary zone electrophoresis for haemoglobinopathy diagnosis

AimsCapillary zone electrophoresis (CE) at alkaline pH is increasingly used in haemoglobinopathy diagnosis. We report our evaluation of automated CE, using the Capillarys 2 Flex Piercing instrument, as a routine diagnostic method for the detection of variant haemoglobins and the diagnosis of β thalassaemia.MethodsA Capillarys 2 Flex Piercing instrument with Phoresis software was evaluated in our laboratory over a 6-week period, comparisons being made with high performance liquid chromatography (HPLC) and, for haemoglobin A2 quantification, with microcolumn chromatography.ResultsThe instrument was easy to use and was suitable for the quantification of haemoglobin A2. Quantification of A2 was precise and the percentage was stable with ageing of the blood specimen. Results differ among HPLC, CE and microcolumn chromatography and use of an instrument-specific, method-specific reference range is therefore recommended until such time as there is standardisation between methods and manufacturers. Common variant haemoglobins were provisionally identified without difficulty. There are some uncommon variant haemoglobins that are detected by HPLC but not by capillary electrophoresis, but the reverse also occurs.ConclusionsCapillary electrophoresis using a Capillarys 2 Flex Piercing instrument is suitable for haemoglobinopathy diagnosis.