Utilizing Venturi effect for automated high-throughput droplet-MS from well plates

High-throughput screening is important in a diverse array of applications including drug discovery, synthetic reaction development, and enzyme engineering. Well plates are often used for sample preparation and containment in such applications, so analytical methods that are compatible with this format are required. Mass spectrometry (MS) is an attractive analytical technique for high-throughput analysis due to its potential for rapid, sensitive, selective, and label-free multiplexed measurements. Here, we present a method that uses the Venturi effect to withdraw droplet samples from a well plate and infuse them to the electrospray ionization (ESI) source of a mass spectrometer. The Venturi effect is generated by flow of nebulizing gas through a constriction at the outlet of the ESI source. The resulting negative pressure allows sample to be pulled to the ESI source via a sample transfer capillary that is coaxial with the ESI source at the outlet and can be dipped into sample at the inlet. To keep different samples from mixing, 380 nL sample plugs flowing at 330 µL min^-1 are sipped into the source and separated by air gaps resulting in segmented flow to the source. The system requires no valves or connections for achieving sample pick-up and analysis. An x,y,z-positioner is used to move the sample inlet for automated sampling from different wells. Increasing capillary inner diameter and nebulizing gas pressure increased the throughput of Venturi droplet-MS by increasing sample flow rate. An interaction of sample plug size and number within the capillary on overall flow rate was observed and affected the possible throughput. When using perfluoroalkoxy alkane (PFA) tubing as the transfer capillary, carryover between samples was 0.88 ± 0.16%. The method is demonstrated by screening 283 whole cell reactions for enzyme engineering at 0.4 samples per s, while showing good agreement (R² = 0.92) with liquid chromatography-mass spectrometry (LC-MS). This work improves upon previous uses of segmented flow for high-throughput MS by integrating sample generation and transfer in one step. Compared to other high-throughput MS methods this approach requires no custom MS sources or specialty sample introduction equipment.