The Diagnosis of α-Thalassaemia: A Case of Hemoglobin H −α3.7 Deletion

Abstract

We report a case of hemolytic anemia that was subsequently identified to be a case of α-thalassaemia harboring the common rightward 3.7 kb deletion/HbH. The diagnosis was based on sequential analyses using BioRad D10 HPLC, Alkaline gel electrophoresis, GPO α THAL-IC strips and the identification of the specific genetic lesion using an α Globin reverse dot blot hybridization assay. Supravital stain of RBCs helped in identifying classical HbH inclusions. In a background of a variable clinical presentation, lack of definitive hematological markers, and general under-diagnosis of α-thalassaemias we have used this case to highlight the features and sequence of techniques involved in identifying and characterizing an α-globin chain mutation, starting from a diffuse clinical history and presentation up to the identification of a specific genetic lesion involved.

Cellular function reinstitution of offspring red blood cells cloned from the sickle cell disease patient blood post CRISPR genome editing

“…To investigate changes in cellular protein expression, the cloned cells were lysed, and hemoglobin types were assessed by a clinically viable HPLC method (8.3 × 10 3 cells/test) as described in “ Materials ” and “ Methods ” (Additional file 1 : Figure S15) [ 30 ]. Due to genome editing resulting in heterozygous HBB*/HbS , a new peak for HbA* protein was detected in the cloned erythroid progenitor cells at a similar level to residual HbS protein (Fig.…”

“…For high-performance liquid chromatography (HPLC) analysis, cell lysates (in 2.0-mL tubes) were loaded on a BioRad D10 HPLC instrument and analyzed [30], following standard clinical protocol in our department. For each test, 0.25 mL of erythroid progenitor cell lysates (~8300 cells/assay) was used.…”

“…Half of the amplified erythroid progenitor cells (5 × 10 4 ) from individual single colonies were incubated in 1.5 mL of D-10 Wash/Diluent Solution (BioRad, Hercules, CA) for 5 min at room temperature. For high-performance liquid chromatography (HPLC) analysis, cell lysates (in 2.0-mL tubes) were loaded on a BioRad D10 HPLC instrument and analyzed [ 30 ], following standard clinical protocol in our department. For each test, 0.25 mL of erythroid progenitor cell lysates (~8300 cells/assay) was used.…”

Hemoglobin J in a patient with severe anemia, a case report from Nepal

“…CE-HPLC complemented by electrophoresis, is the method of choice for characterizing various hemoglobin variants including Hb J(3, 4). Hb J presents as elevated P3 peak on HPLC while thalassemia is detected by the presence of eluted proteins at the retention time between 0 and 1 minutes [ 16 ]. P3 peak up to 6% is considered normal, values 6%–12% indicates suboptimal specimen and values greater than 15% indicates Hb J(3).…”

HEMOGLOBIN H DISEASE AND GROWTH: A COMPARATIVE STUDY OF DHbH AND NDHbH PATIENTS

“…Other non-deletional types include Hb Quang Sze (α2 codon 125 CTG>CCG), Hb Paksé (α2 codon 142 TAA>TAT), Hb Q Thailand (α2 codon 34 GAC>CAC), Hb Saun Dok (α2 codon 109 CTG>CGG), α2 codon 59 (GGC>GAC), α2 codon 0 Δ1bp (-T), α2 codon 30 Δ3bp (-GAG), and α2 codon 35 (TCC>CCC). [4][5][6] The prevalence ratio of DHbH to NDHbH is varied. Although DHbH was found to be more prevalent in several studies, 3,7,8 NDHbH was more prevalent in some studies from Thailand.…”