On the Use of Image Analysis for Hematocrit Evaluation in Dried Blood Spots

Dried blood spots (DBSs) are formed by collecting a small sample of blood on specialized filter paper and allowing it to dry naturally. Various domains of life sciences and drug research extensively use DBSs as a sampling technique. The “Hematocrit (Ht) effect” affects assay bias, and several strategies have been put forth to deal with it, including the correction of quantified concentrations using an appropriate correction factor. The approach was previously applied, following the utilization of an image processing algorithm developed in Matlab® to derive a reliable equation correlating DBS areas to Ht% values. The present work looks more closely at the application of image analysis to the evaluation of Ht in DBS samples. Utilizing image analysis software, DBS samples with known Ht values were processed. Preparation of cards has followed a previously developed protocol for the appropriate formation of uniform area DBSs, irrespective of Ht. The resulting areas showed close resemblance to the respective theoretical areas calculated by applying the correlation equation. Following that, the equation was utilized to determine the Ht values for each sample, and a comprehensive comparison of measured versus calculated Ht was carried out using various statistical approaches for method comparison. The results demonstrated a strong correlation, suggesting the method’s viability in estimating Ht for unknown DBS samples.