Collection and Storage of Capillary Blood in Glass Fibre Filters for Glycated Haemoglobin Measurement by a Microcolorimetric Method

Abstract

SUMMARY.A filter method for collection and storage of capillary blood spots for glycated haemoglobin (gHb) has been developed. Glass fibre filters (GFB) impregnated with 0.8 M boric acid were used to collect and store capillary blood. Haemoglobin from the dried blood spots was eluted into water and determined by Drabkin's method, while gHb in the eluates was determined by the microcolorimetric method. The intraassay coefficients of variation (CVs) were 4.5, 4 -5 and 3.1% at 882, 1101 and 1704 pmol HMF/mg Hb, respectively. The corresponding inter-assay CVs were 8 -6, 8.6 and 6-3%, respectively. A total of 63 paired capillary and venous blood samples were measured by both the direct and GFB method. The GFB method showed excellent correlation with the direct method (r= 0.948 and r= 0.994) after 7 and 14 days' storage at room temperature. The GFB method will enable prior collection and postage of blood samples by patients. Additional key phrases: method comparison; stability study; diabetes mellitusThe measurement of glycated haemoglobin (gHb) provides a useful index for assessment of longterm blood glucose control in diabetic patients since its level reflects the average blood glucose concentration of the preceding 2-3 months.'-4 Most of the presently available methods for gHb5-' are more complex and time-consuming than the simple dry chemistry methods for blood glucose, thus requiring the use of filter paper methods for advanced collection and despatch of blood samples by patients to the laboratory, to ensure that gHb results are available at clinic visits.Several filter paper methods for gHb have been reported which employ either affinity or ionexchange chromatography or c o l~r i m e t r y .~-~ In addition to gHb, glycated plasma proteins are also measured by the filter paper colorimetric methodsY-l2 as there is no prior separation of plasma from red blood cells. The colorimetric method is a robust, precise and cheap method which has been recommended for use in developing ~0untries.I~ We previously used the microcolorimetric method for washed red blood cells spottedonto filter papery but this method is not suitable for direct blood collection by patients. Therefore, we have introduced the use of glass fibre filters which permit direct collection and simultaneous separation of capillary blood. This paper describes the validation and use of the GFB method for gHb measurement. MATERIALS AND METHODSAnalytical reagent grade orthophosphoric acid, and boric acid (Merck), trichloroacetic acid, thiobarbituric acid and 5-hydroxymethylfurfural (HMF; Sigma Chemicals) and glass fibre filters (GF/B; Whatman), were purchased from Chemopharm Sdn. Bhd., 20 Jalan SS2/66,47300 Petaling Jaya, Selangor.A total of 63 paired venous and capillary blood samples were collected from 21 normal staff and 42 diabetic patients of the Medical Faculty, Universiti Kebangsaan Malaysia. Venous blood samples (2 mL) collected in heparanized tubes were centrifuged and the packed red blood cells rinsed twice with normal saline to remove free glucose b...

Prevalence of NIDDM and Impaired Glucose Tolerance in Aborigines and Malays in Malaysia and Their Relationship to Sociodemographic, Health, and Nutritional Factors

“…HbA x was measured by using microcolorimetric method with filter paper blood spots (9). All the subjects selected were invited to attend the clinic, which was set up specifically for the survey.…”