Determination of hemoglobin content in whole blood and in dried blood spots in lambs

Abstract The use of filter paper for the collection and analysis of human blood has begun in the 1960s. In those days Dr. Robert Guthrie used dried blood spot (DBS) specimens to measure phenylalanine in newborns. DBS are whole blood samples collected from blood obtained by finger-sticks or heel pricks and placed onto filter paper, dried and eluted later for laboratory analyses. The purpose of this study was to develop and evaluate a capillary filter paper method that could be employed as a reliable technique for hemoglobin (Hb) determination in animals. Heparinised blood (10 ml) was taken from healthy lambs (n=20). DBS were prepared as venous whole blood samples were aliquoted (20 L) onto Whatman 903 (Schleicher & Schuell GmbH Inc., Germany) filter paper. Either the entire DBS were cut from the paper card with scissors for extraction, or DBS punches were removed from each DBS by using a standard paper hole punch. The discs were placed into test tubes for elution in Drabkin reagent and the eluates were measured spectrophotometrically. Hb values obtained on filter paper were compared with values from fresh whole blood samples. As our results indicate the standard punches gave identical results as the whole blood determinations under similar conditions (2.5 ml reagent and 20 min elution time). After 10 day storage period the elution required more time (40 or 60 min) for the same results. The use of DBS offers certain advantage over using traditional methods and conventional samples. Not only in case of infants but also in animal investigations the DBS are a promising alternative to venipuncture.