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J Biomol Screen. 2016 Jul;21(6):620-5. doi: 10.1177/1087057116630706. Epub 2016 Feb 22.

A Comparison of LC-MS/MS and a Fully Integrated Autosampler/Solid-Phase Extraction System for the Analysis of Protein Binding Samples.

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Drug Metabolism & Pharmacokinetics, Biogen, Inc., Cambridge, MA, USA.
Analytical Sciences & Imaging, Novartis Institutes for Biomedical Research, Inc., Cambridge, MA, USA
Drug Metabolism & Pharmacokinetics, Agios Pharmaceuticals, Cambridge, MA 02139 USA.
Drug Metabolism & Pharmacokinetics, Novartis Institutes for Biomedical Research, Inc., East Hanover, NJ, USA.


A new analysis approach was evaluated for measuring plasma protein binding (PPB) of small molecules using the Agilent RapidFire high-throughput system coupled with a Sciex API 4000 mass spectrometer (RF-MS/MS). Thirty-three proprietary and 12 literature compounds were subjected to rapid equilibrium dialysis (RED) and evaluated in parallel using RF-MS/MS at 16.4 s/sample and traditional liquid chromatography-tandem mass spectrometry (LC-MS/MS) at 3.5 min/sample, thus making the RF-MS/MS analysis over 12 times faster than LC-MS/MS. The high-throughput analysis method that was developed demonstrated excellent correlation with the traditional LC-MS/MS analysis method with an r(2) value of 0.96. The RF-MS/MS analysis method was implemented to increase sample throughput, decrease turnaround time for PPB data, and decrease time burden on existing LC-MS/MS instruments.


RapidFire; high throughput; mass spectrometry; protein binding

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