Bioanalytical sample preparation methods range from simple techniques such as dilute and shoot or protein precipitation to more specialized techniques such as liquid-liquid extraction, solid phase extraction (SPE) or immunoaffinity purification. One of the challenges of bioanalytical sample preparation in modern laboratories is achieving consistent results from batch-to-batch. Technique dependent variables such as pipetting consistency and other user dependent variables can impact accuracy and repeatability. Automating the sample preparation workflow can minimize this variability while freeing up scientists for other tasks.
In this work, the Andrew+™ Pipetting Robot configured with the Extraction+ connected device was used to fully automate sample preparation and extraction of the therapeutic drug, apixaban (Eliquis) from plasma. Several common bioanalytical techniques were employed including protein precipitation (PPT), PPT with phospholipid (PL) removal, supported liquid extraction (SLE), revered-phase (RP) SPE, RP-SPE with PL removal and mixed-mode SPE. Each method including all pipetting steps, vacuum settings, waste disposal and final collection was fully automated. Analyte recovery, matrix effects and residual PLs were used to compare efficiency and cleanliness of the methods.
Across all extraction techniques, recoveries ranged from 53-100% with all but one (SLE) >75%. Matrix effects varied based on the technique ranging from -41% to 35%. Residual phospholipids were highest for PPT and lowest for SLE, and generally decreased with increased method specificity (RP-SPE>PL removal>mixed-mode SPE). Across all techniques, quantitative performance was excellent, with sample accuracies within 10% of target values and %RSDs in the single digits for all QC levels. Calibration curves were linear and accurate across the dynamic range (2-500 ng/mL) with accuracies ranging from 86-111% and % RSDs between 0.2-15%. As with the QC results, most were in the low single digits. These criteria easily meet regulatory guidances for bioanalytical method performance.
This work demonstrates the capability of the Andrew+ Pipetting Robot with Extraction+ connected device to reliable execute a diversity of sample extraction techniques-simple to complex-used in bioanalysis. The consistency and reliability of the platform was demonstrated by the excellent accuracies and precisions achieved, with most in the low single digits. This combination of the Andrew+ Pipetting Robot with and Extraction+ connected device provides an easy to use, accessible solution for automating bioanalytical sample preparation improving method reliability and freeing up scientists for other tasks.