A rare haemoglobin-variant produces discrepant results between haemoglobinopathy screening techniques

Abstract

The validity of a haemoglobinopathy diagnostic technique is dependent on several factors including the sensitivity and specificity of the methodology used. We describe a case of a 69-year-old man who was referred for investigation of a chronic, mild normocytic anaemia. Initial investigations by a private, interstate pathology service reported a fast-migrating haemoglobin-variant of over 40% in Zone 11 using a capillary zone electrophoresis technique (Sebia Capillarys). However, confirmatory testing in a major haemoglobinopathy screening centre failed to identify the presence of a haemoglobin-variant using a high performance liquid chromatography technique (HPLC; Biorad Variant II) in conjunction with agarose gel at acid pH and cellulose acetate at alkaline pH haemoglobin electrophoresis techniques. Extensive investigation of pre-analytic variables and laboratory variables ensured the accuracy of the discrepant results reported by both laboratories. Subsequent molecular sequencing identified the pathognomonic amino acid substitution of Arginine (AGG) for Serine (AGT) at codon 30 of the β haemoglobin gene consistent with haemoglobin Tacoma. With nearly 70% of the laboratories participating in the RCPA 2013 Haemoglobinopathy Program using HPLC techniques, this case suggests that some haemoglobin-variants will remain undiagnosed due to limitations in the sensitivity of the methodology used. The validity of a haemoglobinopathy diagnostic technique is dependent on several factors including the sensitivity and specificity of the methodology used. We describe a case of a 69-year-old man who was referred for investigation of a chronic, mild normocytic anaemia. Initial investigations by a private, interstate pathology service reported a fast-migrating haemoglobin-variant of over 40% in Zone 11 using a capillary zone electrophoresis technique (Sebia Capillarys). However, confirmatory testing in a major haemoglobinopathy screening centre failed to identify the presence of a haemoglobin-variant using a high performance liquid chromatography technique (HPLC; Biorad Variant II) in conjunction with agarose gel at acid pH and cellulose acetate at alkaline pH haemoglobin electrophoresis techniques. Extensive investigation of pre-analytic variables and laboratory variables ensured the accuracy of the discrepant results reported by both laboratories. Subsequent molecular sequencing identified the pathognomonic amino acid substitution of Arginine (AGG) for Serine (AGT) at codon 30 of the β haemoglobin gene consistent with haemoglobin Tacoma. With nearly 70% of the laboratories participating in the RCPA 2013 Haemoglobinopathy Program using HPLC techniques, this case suggests that some haemoglobin-variants will remain undiagnosed due to limitations in the sensitivity of the methodology used.