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November 15, 2009 (Vol. 29, No. 20)

Dried Blood Spots for Toxicokinetic Studies

Reduction of Sample Volumes Required Is One of the Key Benefits of DBS Assays

  • Assessment of Performance

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    Figure 2. The accuracy of the dried blood spot method for different blood volumes for acetaminophen in dog blood

    Accuracy and precision are vital elements in a TK study, and the impact of using DBS has been assessed using quality control samples at four different concentrations for both dextromethorphan (DM) and dextrophan (DX). These samples were analyzed against an LC-MS/MS calibration curve, constructed using spiked DBS standards.

    The accuracy, defined as the percent deviation from the theoretical concentration, was generally within 10% for all the quality control samples. Relative standard deviation (RSD) for the replicate analyses, where n=6, was typically less than 10%. The conclusion from this data is that DBS provides the accuracy and precision required for TK studies.

    In addition, two other factors that could potentially affect the quality of the resultant data were investigated—volume of blood sample applied to the card and consistency with the blood spot.

    It has been found that, although a consistent blood volume can be helpful for generating more consistent results, accurate pipetting of the sample onto DBS cards is not required. When QC samples of three different concentrations of acetaminophen were taken at volumes of 10, 20, 40, and 50 µL, a small increase in accuracy was observed in only the larger 40 and 50 µL volumes (Figure 2). Therefore, it was concluded that full GLP calibration was unnecessary. Thus DBS provides a simple blood-sampling procedure that can be easily used at the site of sampling.

    To assess the effect of punch position within a blood spot, QC samples punched from the center of a larger blood spot were compared to those from closer to the edge of the spot. Any variations fell within the measurement error and can be considered as having good precision with no significant difference. The results support analyte concentration homogeneity across a blood spot made with good technique on a good matrix.

  • Summary

    The DBS assay has been shown to be a viable method for performing TK analyses, displaying the necessary accuracy and precision for such experiments. The reduction in the required volume of blood enables collection of more samples from each test animal, thereby directly reducing the number of animals while at the same time increasing the quality of the data generated.

    TK studies using DBS have the added benefit that samples can be stored and safely shipped at ambient temperatures. Using DBS not only increases the speed of testing by reducing sample processing steps, but can also contribute cost savings in preclinical TK testing. 

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