Comparison of fast protein liquid chromatography (FPLC) with HPLC, electrophoresis & microcolumn chromatography techniques for the diagnosis of beta-thalassaemia.

BACKGROUND & OBJECTIVE: beta-thalassaemia is a genetic disorder and an important health problem around the world. Quantitative haemoglobin A(2) (HbA(2)) levels are used for the diagnosis of beta-thalassaemia. The conventional methods are high performance liquid chromatography (HPLC), electrophoresis, and microcolumn chromatography techniques. We established a fast protein liquid chromatography (FPLC) method, to measure quantitatively of HbA(2) levels, and compared its efficacy with conventional methods. METHODS: The FPLC method, using a DEAE Sepharose, Hi Trap anion-exchange column chromatography technique was set up for HbA(2) measurement. In this study, 220 blood samples were screened for haemoglobin type by FPLC technique and also using HPLC, microcolumn chromatography and electrophoresis. RESULTS: The FPLC results were highly correlated (r = 0.985, P<0.001) with those of HPLC for quantification of HbA(2) as well as cellulose acetate electrophoresis (r = 0.977) and microcolumn chromatography (r = 0.980). The FPLC method showed 100 per cent sensitivity and specificity, positive and negative predictive value for beta-thalassaemia diagnosis. In addition, the FPLC method was simple, rapid, low cost and reproducible. The HbA(2)/E range of FPLC for beta-thalassaemia was 6-10 per cent, HbE trait was 10-40 per cent, beta-thalassaemia/HbE was 40-60 per cent and homozygous HbE was more than 60 per cent. INTERPRETATION & CONCLUSION: Our findings suggested that FPLC method could be used as a cost-effective method for routine beta-thalassaemia diagnosis.

Evaluation of a random access analyser: BM/Hitachi 911.

The performance of Boehringer Mannheim's BM/Hitachi 911 was evaluated for three months. The mean coeffcient of variation (CV) of the within-run and between-run imprecision of the 16 analytes were less than 1.16% (range 0.47-2.38%) and 1.35% (range 0.62-2.93,%), respectively. A linearity study for the various assays covered clinically important levels. No relevant drift was observed during an eight-hour assay nor was any sample-related carry-over detected. In all cases, the regression analyses (slopes) of the results obtainedfrom BM/Hitachi 911 and 717 were between the extreme values of 0.94 and 1.05. During the three months of operation, no major problem was encountered. The BM/Hitachi 911 was found to be easily operated, to require minimal attention and simple daily maintenance during operation.

Laboratory evaluation of the COBAS MIRA S random access analyser.

The random access analyser COBAS MIRA S (Roche Diagnostics) was evaluated for two months. The instrument is a computer-controlled discrete analyser which can be run in a combination profile and single test mode. This instrument has special features, including an automatic cuvette segment changer, a reagent rack cooling system, an external keyboard and monitor, as well as a bar-code facility for the entry of test parameters, worklists and sample identification numbers. Study of within-run and between-run precision gave values of % CV 0.54-3.37 and 0.61-3.65, respectively, for a variety of assays. Linearity testing to the upper limit of each test was also studied and were found to cover the necessary pathological range. Within the two-month period, no major problems were encountered. The instrument required minimum operator attention during operation. Correlation studies with the Hitachi 705 using six clinical chemistry tests (glucose, cholesterol, triglyceride, ALP, AST, ALT) gave correlation coefficients ranging from 0.95-0.99 and slopes of 0.91-1.17.