Figure 3

Chemical pathways for Flyspresso. (A) The System Piston pushes and pulls the System Liquid (blue) to either the Chemical Syringe Pumps or the Priming Syringe Pump. A solenoid valve controlled by the Arduino board, automatically switches between the vacuum and gas supply to move the piston. (B) The Flyspresso robot is equipped with six Chemical Syringe Pumps. A controllable solenoid valve opens and closes to allow the system liquid to move each Chemical Syringe Pump. The chemical input (orange) is pulled into the syringe and pushed into each corresponding well. Passive check-valves (small rectangles with circles inside) prevent chemicals from flowing in the opposite direction. Chemical waste is also aspirated from the wells (magenta), but does not flow back through the Chemical Syringe Pump. (C) The Priming Syringe Pump clears the tubing when a new chemical is used. Similar to the Chemical Syringe Pumps, the Priming Syringe Pump is controlled by a solenoid valve and displacement of the System Liquid. (D) Chemical waste from aspiration and priming is carried to waste containers. The waste containers are connected to a solenoid valve, which moves to open or close the vacuum. An activated charcoal filter prevents volatile fumes from being pulled into the vacuum. The entire system is enclosed in a chemical fume hood. (E) The Chemical Manifold is a modular and expandable set of solenoid valves attached to reagent bottles. The solenoid valves alternate between ambient air and the vacuum. When not in use, the vacuum prevents the chemical from being pulled into the syringes. When in use, the solenoid valve switches to ambient air, allowing the chemical to pass through a one-way passive check-valve. This valve prevents the chemicals from flowing into and contaminating other containers. (F) Cut-through sectioning of the microplate showing the syringe tips for the Chemical Syringe Pumps (orange) and separation tips (blue) from the Separation Manifold (see also Fig. 2). The Transplate (24W TRANSPLATE.ipt) rests on the base of the microplate. Embryos are depicted in magenta and rest in the individual meshes of the Transplates. The Seplate attachment (24 SEPLATE.ipt) inserts above the Transplate and contains inverted cones with small openings for removing chemicals with the Separation Manifold tips. (G) (Left) Chemicals from the Chemical Syringe Pumps are loaded into the microplate wells. The tips from the syringes can also aspirate chemicals to chemical waste (magenta). (Middle) During isotonic shocking, embryos that fail to separate and vitelline membranes float to the surface through the Seplate Attachments. The Seplate Attachments nest in the Transplates and contain an inverted cone with a small, one-way orifice. Shocking forces floating membranes and debris through the small, one-way orifice, while successfully shocked embryos do not float and are protected from being aspirated by the Seplate Attachments. (Right) The tips from the Separation Manifold (blue) remove chemicals from this upper surface without disturbing successfully separated embryos at the base of the Transplate (white).