Photomultiplier tube detector

Excitation was provided by an LED source (X-Cite) operated at minimum (5%) output at 505–545 nm through a 20×/0.46 NA objective (Olympus). The illumination intensity was further reduced using a 0.4 ND (Semrock) filter to minimize photobleaching and filtered through a 525/50-nm bandpass filter (Semrock). A 562-nm dichroic (Semrock) and a 593/46-nm bandpass filter (Semrock) were used to separate fluorescence emission from reflected light. Fluorescence signals were detected with a Hamamatsu photomultiplier tube detector and transmitted directly to the Digidata 1440 A A/D converter at a frequency of 10 kHz. Oocytes were recorded from a 0-mV holding potential, or −60-mV holding potential with a −110-mV prepulse for 500 ms in paired recordings as was done for gating current recordings. Interpulse duration was 2 min and test pulse duration was 25 s for all recordings.

A custom‐built single‐laser, single‐color fluorescence detection system was designed for electrophoretic separation and laser‐induced fluorescence (LIF) detection (Figure S2). The microdevice is placed on a metal stage that can be heated for electrophoresis and adjusted in three dimensions to focus the incident laser through a hole in the stage into the middle of the separation channel of the microfluidic device. Voltage‐driven electrophoresis was achieved by placing platinum electrodes connected to a high‐voltage power supply in the 3D printed reservoirs on the microfluidic chip seated on the metal stage. The custom‐built voltage supply system is controlled by LabVIEW. The fluorescence detection system consisted of a 488‐nm solid‐state laser for excitation, which was directed to a mirror, through a 525‐nm short pass dichroic mirror, and into the end of a 40× LD Acroplan, 0.6 NA objective (Zeiss, Thornwood, NY, USA) where it focused into the microdevice separation channel. The emission beam was passed back through the objective, reflected off the dichroic mirror, and passed through a 505‐nm long pass filter toward a photomultiplier tube detector (Hamamatsu, Tokyo, Japan). Data collection and electrophoresis conditions were controlled by a custom‐made LabVIEW program.