BACKGROUND:More than 900 hemoglobin (Hb) variants are currently known. Common techniques used in Hb analysis are electrophoretic and chromatographic assays. In our laboratory, we routinely apply chromatographic methods. To ascertain whether Hb variants are missed with our procedures, we additionally analyzed all samples with mass spectrometry (MS).
A new alpha-globin mutation [alpha 138(H21)Ser-->Phe] was found in a 55-year-old male proband with an erythrocytosis known since his youth. Cation exchange high performance liquid chromatography (HPLC) revealed an additional peak eluting slightly before Hb A indicating the presence of a variant. The peak area of the variant was approximately one-third that of Hb A suggesting an alpha-globin variant. Matrix-assisted laser desorption ionization-time-of-flight mass spectrometry analysis confirmed the mutation at the protein level. The variant is also detectable with isoelectric focusing and reversed phase HPLC. DNA analysis revealed a heterozygous sequence mutation at codon 138 of the alpha2 gene. A C>T transition at the second nucleotide of the codon indicated a Ser-->Phe exchange. The variant showed increased oxygen affinity and was named Hb Frauenfeld.
A 14-year-old male child presented with microcytosis, a known alpha(+)-thalassemia (alpha-thal) heterozygote and a hemizygous glucose-6-phosphate dehydrogenase (G6PD) deficiency. Furthermore, cation exchange high performance liquid chromatography (HPLC) revealed an additional peak eluting slightly before Hb A. The peak area of the variant was equal to that of Hb A, suggesting a beta-globin variant. Matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS) analysis confirmed the mutation at the protein level. The variant was detectable by isoelectric focusing (IEF) or by reversed phase HPLC. DNA sequencing revealed a heterozygous mutation at codon 135 of the beta gene, already described as Hb Alperton. Hb Alperton showed decreased oxygen affinity. Neither biochemical nor clinical characteristics for Hb Alperton have been reported so far.
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