False positive rates of thalassemia screening in rural clinical setting: 10-year experience in Thailand.

This retrospective study aimed to describe the magnitude of false positive screening for thalassemia in a primary care setting of Thailand. The study was conducted from 1999 to 2008 and analyzed 13,745 positive cases. It involved a combination of one tube osmotic fragility (OF) and dichlorophenol indophenol (DCIP) precipitation tests. The number of cases increased over the ten-year period, corresponding well to an exponential model. Based on hemoglobin and DNA analysis, cases with alpha-thalassemia 1, beta-thalassemia, and hemoglobin E were defined as true positive cases, and the remaining were considered as false positives. The false positive rate was in the range of 20.1-36.1%. The proportion of false positive cases of thalassemia from the screening tests was associated with a trend which was statistically significant (p < 0.001). The estimated cost of further hemoglobin analysis resulting from false positive cases was approximately 40.2-72.2 million THB/year for an estimated 800,000 annual births. The combination of the OF and DCIP test, which has been the strategy for screening of thalassemia and HbE in pregnant women throughout this country, resulted in a large economic burden in terms of high cost and workload associated with further hemoglobin and DNA analyses of false positive samples. Measures to reduce false positive should be developed and implemented.

A spurious haemoglobin A(1c) result associated with double heterozygote for haemoglobin Raleigh (ß1[NA1]Val → Ala) and α(+)-thalassaemia.

BACKGROUND: Accurate measurement of haemoglobin A(1c) (HbA(1c)) is useful for long-term glycaemic control in patients with diabetes. Many Hb variants can interfere with HbA(1c) measurement and cause inaccurate results. METHODS: The subject was a 31-year-old Thai man who was discovered because of an unexpected HbA(1c) result; other diabetic parameters were within the normal range. Abnormal Hb was investigated using automated high-pressure liquid chromatography (HPLC) and a capillary electrophoresis system. Mutation analysis was done by cDNA sequencing, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and multiplex allele-specific PCR assays. RESULTS: Evaluation of HbA(1c) by cation-exchange HPLC showed a value of 34.9% (reference interval, 4.0-6.0%), but a value of only 4.0% (reference value, 4.8-5.9%) was found with a turbidimetric immunoassay. Haematological analysis revealed a mild anaemia but other parameters were within the normal range. Hb-HPLC analysis demonstrated an unknown Hb variant (47.0%) separating from HbA (46.7%), but capillary electrophoresis identified no abnormal peaks. Mutation analysis identified the Hb Raleigh (ß1[NA1]Val → Ala [GTG → GCG]) mutation in combination with an α(+)-thalassaemia, a hitherto undescribed association. The Hb Raleigh mutation could be detected by PCR-RFLP or a multiplex allele-specific PCR assay. CONCLUSIONS: Hb Raleigh can cause falsely increased HbA(1c) values on cation-exchange HPLC. Definitive diagnosis of this variant using combined Hb and DNA analyses is therefore essential.