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Evaluation of a new semi-automated high-performance liquid chromatography method for glycosylated haemoglobins

Hindawi Publishing Corporation
Publication Date
DOI: 10.1155/s1463924686000378
  • Research Article
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Journal of Automatic Chemistry of Clinical Laboratory Automation, Vol. 8, No. 4 (October-December 1986), pp. 192-196 Evaluation of a new semi-automated high-performance liquid chromatography method glycosylated haemoglobins Assunta Carpinelli, Andrea Mosca* Universitd degli Studi di Milano, Dipartimento di Scienze Tecnologie Biomediche, Via O{gettina 60, 1 20132 Milano, Italy and Pierangelo Bonini Istituto Scientifico S. Raffaele Milano, Italy Introduction The measurement of glycosylated haemoglobin in blood is vital for monitoring metabolic control in diabetics [1-3]. The development of many different methods of measurement (based on cation-exchange chromato- graphy, electrophoresis, high performance liquid chro- matography [HPLC], affinity chromatography, radio and colorimetric immuno techniques) proves the impor- tance of this assay [4]. Since the first HPLC method was described [5] a number of modifications have been reported (see the review by Ellis et al. [6]). Instruments have been specially designed and commercialized (for example by Helena Laboratories, Beaumont, Texas, USA and Kyoto Daichi, Japan). The evaluation of a recently commercialized semi- automatic HPLC instrument, which performs analyses on hemolysates from the original blood samples, is reported. Materials and methods The instrument, commercialized by Bio-Rad Labora- tories (Segrate, Milano, Italy), consists of a main unit, containing the functional parts (autosampler, buffer reservoir, pumps, column colorimeter, and a control unit). A single piston pump is used with a step-gradient valve system, by which the elution of the column is performed with three phosphate buffers ofpH 5"7, 5"8 and 5"9 (unstated concentration) and increasing ionic strength. The analytical column (4"0 x 150 mm) is filled by a new cation exchanger (TSK gel, unstated composi- tion). The column temperature is controlled at 23 C by forced ventilation in the column compartment. The sample (5 !1 of venous blood, with mg/ml of EDTA) is diluted automatical

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