1/14. Laboratory recognition of a rare hemoglobinopathy: hemoglobins SS and SG(philadelphia) associated with alpha-thalassemia-2.This article describes the laboratory investigation of an unusual hemoglobinopathy involving hemoglobin (Hb) S, HbSG(philadelphia), and alpha-thalassemia-2 in a patient whose phenotype was HbSC by alkaline electrophoresis. Findings of a mean corpuscular volume of 62 fL and microcytes on the blood smear were inconsistent with HbSC disease. The patient's clinical course over several years had been mildly symptomatic. Testing in our hospital laboratory using isoelectric focusing and cation-exchange high-performance liquid chromatography to separate hemoglobins showed an unknown variant. Additional studies, including globin chain electrophoresis, reverse-phase high-performance liquid chromatography, and polymerase chain reaction-based dna analysis were performed at reference laboratories, which reported the following findings: HbG(philadelphia) associated with alpha-thalassemia-2, HbS and HbG(philadelphia), and the alpha-globin deletions defining the -alpha3.7/-alpha3.7 genotype. The hemoglobin molecular defects, alpha-thalassemia-2, and the pattern of inheritance are discussed.- - - - - - - - - - ranking = 1keywords = chromatography (Clic here for more details about this article) |
2/14. Hb Groene Hart: a new Pro-->Ser amino acid substitution at position 119 of the alpha1-globin chain is associated with a mild alpha-thalassemia phenotype.alpha-thalassemia (thal) is generally considered to be an expression defect caused mostly by deletions silencing one or more alpha-globin genes. Although nondeletional alpha-thalassemia is considered rare, in our laboratory we frequently observe alpha-thal phenotypes induced by point mutations. We report a new point mutation generating an abnormal hemoglobin (Hb) associated with a mild alpha-thal phenotype in two members of a Moroccan family, who presented with mild but persistent microcytic hypochromic parameters and a balanced beta/alpha synthetic ratio. All attempts to separate an abnormal native or denatured fraction were unsuccessful using electrophoresis, isoelectrofocusing (IEE), ion exchange and reversed phase high performance liquid chromatography (HPLC), denaturing polyacrylamide gel electrophoresis (PAGE), and electrospray mass spectrometry (ES/MS). The anomalous protein was only predictable by dna analysis. The mutated gene product, not separable with any of the techniques used, could be a monomer unsuitable for tetramer formation, which is proteolyzed at an early stage. Alternatively, this mutation could perhaps lead to an abnormal splicing. The CCTCT sequence generated by the mutant, not found in the translated region of the gene, but normally present at the end of the IVS-II, could induce a possible exon skipping. This mutant could generate a mild or a critical Hb H disease in combination with one of the common alpha0-thal deletion defects.- - - - - - - - - - ranking = 0.5keywords = chromatography (Clic here for more details about this article) |
3/14. Complex interaction of Hb Hekinan [alpha27(B8) Glu-Asp] and Hb E [beta26(B8) Glu-Lys] with a deletional alpha-thalassemia 1 in a Thai family.Hemoglobin (Hb) Hekinan (alpha27; Glu-Asp) is a rare alpha-chain variant found mainly in Japanese and Chinese whereas Hb E (beta26; Glu-Lys) is common among Southeast Asians. We report a hitherto undescribed condition in which these two variants co-segregate. The proband was a 25-yr-old Thai woman who was encountered with the presence of mild hypochromic microcytosis with Hb 8.2 g/dL, hematocrit (Hct) 26.0%, Mean Corpuscular Value (MCV) 68.6 fL, Mean Corpuscular Hemoglobin (MCH) 21.6 pg and Mean Corpuscular Hemoglobin Concentration (MCHC) 31.5 g/dL. Although Hb electrophoresis at alkaline pH did not show any abnormal band except Hb E in addition to Hb A, high performance liquid chromatography analysis revealed abnormal peaks at the Hb A and Hb E positions. dna analysis of the proband revealed a GAG-GAT mutation at codon 27 of the minor alpha1-globin gene for Hb Hekinan in trans to the South-east Asian (SEA) deletional alpha-thalassemia 1 determinant and a GAG-AAG mutation at codon 26 of the beta-globin gene for Hb E. She was therefore a triple heterozygote for these three anomalies. family study identified that her mother was a double heterozygote for Hb Hekinan and Hb E without alpha-thalassemia whereas her father was a classical Hb H disease patient. The genotype-phenotype relationship observed in this Thai family with complex hemoglobinopathies is presented and a simple dna assay based on the polymerase chain reaction methodology for rapid diagnosis of Hb Hekinan is described.- - - - - - - - - - ranking = 0.5keywords = chromatography (Clic here for more details about this article) |
4/14. Hb Bronte or alpha93(FG5)Val-->Gly: a new unstable variant of the alpha2-globin gene, associated with a mild alpha( )-thalassemia phenotype.We report a new unstable variant identified in three carriers of a family from East sicily; it was named Hb Bronte after the place from which the family originated. dna sequencing from nucleotides -181 to 894 (alpha1) and to 884 (alpha2) revealed a GTG-->GGG substitution at codon 93 of the alpha2-globin gene. The MCV and MCH values were at the lower end of the normal range in the carriers. On cation exchange high performance liquid chromatography (HPLC), the Hb A2 level was apparently increased to around 6%, and a small abnormal peak (0.3-0.4%) was detected after Hb A2. Two abnormal bands were detected by cellulose acetate electrophoresis: a major band (about 3-4%) migrated between Hb A and Hb F; a minor band (<1%) migrated between Hb A2 and carbonic anhydrase. Normal values of Hb A2 were detected by DEAE microchromatography. On reversed phase HPLC the variant chain was not detected, and most likely it was eluted with the alpha chain peak. The isopropanol stability test was very slightly positive in the carriers. Hemolytic symptoms were absent with the exception of indirect bilirubin, which was at high borderline in 2/3 carriers. In biosynthesis in vitro, the specific activity of the alpha chains was much higher than that of the beta-globin chains, and the alpha/beta biosynthetic ratio in the mother and proband was of the beta-thalassemia (thal) type (2.24 and 2.54, respectively). time course experiments showed that the increase of the 3H-specific activity of the peak containing normal and variant alpha chains was not linear and was much higher than that of beta chains; moreover, the alpha/beta biosynthetic ratio varied during the 2 hours incubation.- - - - - - - - - - ranking = 1keywords = chromatography (Clic here for more details about this article) |
5/14. Failure of microchromatographic measurement of fetal hemoglobin in beta zero thalassemia-hereditary persistence of fetal hemoglobin.We report microchromatographic measurement of fetal hemoglobin (HbF) proportions in a 36-year-old African-American multigravida woman. At 34 weeks she delivered a 630-g male infant who subsequently did well. Hemoglobin electrophoresis of the hemolysate revealed nearly 100% HbF without HbA, an extremely unusual naturally occurring sample. family studies revealed a combination of hereditary persistence of fetal hemoglobin (HPFH) and beta zero-thalassemia minor. Southern blot technique confirmed heterozygous alpha 2 thalassemia and HPFH but failed to identify the beta thalassemic lesion. The absence of HbA and the very high amounts of HbF led us to measure HbF by several methods to confirm the accuracy of microchromatography of HbF at values approaching 100%. HPLC revealed a 14% F1 suggestive of microchromatographic underestimation due to glycated HbF. We conclude that cation-exchange microchromatography and the Betke method of alkali denaturation underestimate HbF values as they approach 100% and do not recommend these procedures in this rare situation.- - - - - - - - - - ranking = 1keywords = chromatography (Clic here for more details about this article) |
6/14. Hb Suan-Dok [alpha109(G16)Leu-->Arg; CTG-->CGG (alpha2)] described in a patient of African ancestry.A 58-year-old Black female from Curacao (west indies) was recently referred to our Laboratory for a persistent microcytic hypochromic anemia. An analysis 13 years earlier had shown no abnormal hemoglobin (Hb) fractions and a balanced beta/alpha synthetic ratio. The hematological indices were again compatible with thalassemia and no abnormal fractions were observed on electrophoresis or high-performance liquid chromatography (HPLC). None of the seven common alpha-thalassemia (thal) deletion defects were present. Direct sequencing of the alpha2 gene revealed a CTG-->CGG single base substitution at codon 109. This mutation was previously described in a Thai patient (Hb Suan-Dok), inducing Hb H disease in association with a - -(SEA) allele. In contrast with earlier reports we were unable to identify any native Hb fraction. The balanced beta/alpha ratio indicated that alpha2-Suan-Dok is formed but does not form tetramer formation unless alpha-thal is present.- - - - - - - - - - ranking = 0.5keywords = chromatography (Clic here for more details about this article) |
7/14. Laboratory diagnosis of a compound heterozygosity for Hb Hekinan [alpha27(B8) Glu-Asp] and a deletional alpha-thalassaemia 2 in thailand.We report the haematological and molecular characterization of a previously undescribed condition of compound heterozygosity for haemoglobin (Hb) Hekinan [alpha27(B8) Glu-Asp] and a deletional alpha-thalassaemia 2 detected in a Thai individual. Hb analysis demonstrated that although this Hb variant co-migrates with Hb A on cellulose acetate electrophoresis and cation-exchange high-performance liquid chromatography (HPLC), the HPLC procedure using a weak cation-exchange material with polyaspartic acid could clearly differentiate the two Hb. The variant could then be confirmed using the polymerase chain reaction-restriction fragment-length polymorphism (PCR-RFLP) analysis of the amplified alpha1-globin gene.- - - - - - - - - - ranking = 0.5keywords = chromatography (Clic here for more details about this article) |
8/14. Molecular and hematological characterization of hemoglobin hope/hemoglobin e and hemoglobin hope/alpha-thalassemia 2 in Thai patients.We report the hematological and molecular characterization of compound heterozygosity for hemoglobin (Hb) hope/Hb E and double heterozygosity for Hb hope/ alpha-thalassemia 2 found in 2 unrelated Thai individuals. The first proband presented with slight anemia and mild hypochromic microcytosis. Routine cellulose acetate Hb electrophoresis at pH 8.6 revealed in addition to Hb E another variant migrating slightly more anodic to Hb A. On cation exchange high-pressure liquid chromatography, the variant was eluted in the amount of 60.9% after Hb E The same abnormal Hb was found in a second family in which the proband and her younger sister were both double heterozygotes for this Hb variant and deletional alpha-thalassemia 2, whereas an older sister was a pure carrier of the variant. The amounts of this variant were found to be 34.9%, 35.4%, and 38.3% in the proband, her younger sister, and her older sister, respectively. Direct dna sequencing of the amplified beta-globin genes of both probands identified the GGT (Gly)-GAT (Asp) mutation at codon 136 corresponding to Hb hope. beta-Globin gene haplotype analysis demonstrated that all the Thai betaHope genes were associated with the same haplotype, ( - - - - ), indicating a single origin of this variant in thailand. A simple method based on allele-specific polymerase chain reaction for accurate diagnosis of the Hb hope is described.- - - - - - - - - - ranking = 0.5keywords = chromatography (Clic here for more details about this article) |
9/14. Hb Oegstgeest [alpha104(G11)Cys-->Ser (alpha1)]. A new hemoglobin variant associated with a mild alpha-thalassemia phenotype.A microcytic hypochromic anemic state was observed in an 8-year old Black female of Surinam origin during pre-operative Hb S [beta6(A3)Glu-->Val] screening. Her high zinc protoporphyrin (ZPP) level suggested a chronic iron depletion but, in contrast, the high red blood cell (RBC) count (5.85 x 10(12)/L) was indicative of a possible coexisting thalassemia. No abnormal hemoglobin (Hb) bands were present on high performance liquid chromatography (HPLC) or alkaline electrophoresis and the Hb A2 level was normal. Break point polymerase chain reaction (PCR) failed to reveal any of the common alpha-thalassemia (thal) mutations but selective dna sequencing of both alpha-globin genes disclosed a TGC-->AGC transversion at codon 104 of the alpha1 gene. cystine at codon 104 is involved in alpha/beta globin contact and has been described to be a critical amino acid of the alpha2 chain when substituted by a tyrosine (Hb Sallanches), inducing Hb H (beta4) disease in the homozygous state. Our heterozygous patient had a moderate anemia of 12.2 g/dL and a borderline haptoglobin suggesting some degree of hemolysis.- - - - - - - - - - ranking = 0.5keywords = chromatography (Clic here for more details about this article) |
10/14. Hb Amsterdam [alpha32(B13)Met--Ile (alpha2)]: a new unstable variant associated with an alpha-thalassemia phenotype and a new African polymorphism.We have characterized a new abnormal hemoglobin (Hb) at position 32 of the alpha-globin chain. The proband, a 38-year-old woman of Surinamese Black ancestry, was referred to the Academic Hospital in Amsterdam, The netherlands, after 3 years of prednisone treatment in Surinam. kidney failure was diagnosed at the nephrology Department, Free University Medical Center, Amsterdam, The netherlands; the cortisone treatment was interrupted and dialysis was started. At this stage, a microcytic hypochromic anemia was observed with high reticulocyte (40%) and ferritin (500 microg/L) levels, and hemoglobinopathy was suspected. No abnormal bands were visible on alkaline electrophoresis and high performance liquid chromatography (HPLC). The Hb A2 level was normal (2.7%) and the erythrocyte count was low (3.59 x 10(12)/L) with a normal haptoglobin level (68 mg/100 mL). None of the common alpha-thalassemia (thal) deletion defects were present. The beta-globin gene sequence was normal but the alpha2-globin gene sequence revealed an ATG-->ATA transition at codon 32, changing the methionine into an isoleucine residue. The mutation, called Hb Amsterdam, was observed in the mother of the proband, who was also heterozygous for the--alpha3.7-thal deletion and affected by a moderate microcytic hypochromic anemia. Both Hb Amsterdam and the--alpha(-3.7) allele were found in association with a new polymorphism, IVS-I-39 (C-->T), previously observed in our laboratory in seven patients of African origin, on both the alpha1 and alpha2 genes. In addition, Hb Amsterdam was also associated with the common African alpha2 polymorphism (G-->CTCGGCCC at position 7238 and T-->G at position 7174). Hb Amsterdam is the first mutation ever described at codon alpha32, a position involved in alpha1/beta1 interaction. The possibility of a contribution of this mutation to the nephropatic state of the proband is discussed.- - - - - - - - - - ranking = 0.5keywords = chromatography (Clic here for more details about this article) |
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