Table 1.
Open-source microinjection devices described in the literature.
Fig 1.
Basic design and function of a portable self-contained, externally-actuated microinjector.
A. Overall electronics block diagram of the injector module. The device is powered by a 5V, 3A external AC-DC switching power supply, which supplies all control, pressurization, and airflow. Motor speed and therefore pump pressure is controlled by a simple, off-the-shelf PWM Module utilizing a NE555 timer to generate a dial-controllable pulse train feeding a P2003BDG or LR7843 N-channel MOSFET. The result is high-frequency, low-side PWM of the pump motor, with high-side motor control by a foot pedal or other external switch. Direction control of airflow (aspirate or inject mode) is managed by a pair of solenoid valves that are separately switched, on for aspirate, off for dispense. Note the solenoids themselves are high-side switched on/off with the footswitch as well, as empiric testing found this configuration was necessary to release negative pressure built up in the tubing (when needle filling is completed), which would otherwise be transmitted to the injection needle (creating continued aspiration) even after the high-side switch was released. B. Tubing schematic showing the unidirectional mini air pump, which is connected to two three-way mini solenoid valves that control airflow direction to the microinjection needle. C. In aspirate mode, solenoids are actuated, closing the bottom metal ports and opening the top plastic ports, a configuration that results in the pump inflow port being connected to the microinjection needle. In inject/ dispense mode, solenoids are not powered, closing the top plastic ports of the valves and opening the bottom metal ports, which causes the pump outflow port to be connected to the microinjection needle. D. Wiring diagram of the main microinjector, for user assembly. Detailed parts list is available in S1 Table.
Fig 2.
Completed microinjector and needle holder options.
A. The completed microinjector fits ($85, assembly described below) into a 115 x 90 x 55 mm project box, with a USB-C power connection, a 2-pin GX16 aviator port for pump control, and a 2 mm hose barb fitting for pressure output to microinjection needle. Built-in controls include a injection/aspiration potentiometer knob, which controls motor PWM (aspirate/inject pressure), as well as a SPST toggle switch that controls solenoid valves for airflow direction (aspirate or inject). Optional pulse control module ($75, requires additional assembly, see Fig 7 and 8) also fits into a second similar package, and interfaces with the main injector via its footswitch GX16 port. B. Needle holder options include Word Precision Instruments 5430-ALL ($165 part), Tritech Research MINJ-4 ($78 total, not tested here), or custom home-built needle holder ($36 total, assembly described below).
Fig 3.
Assembly of the microinjector.
A. All parts of the main injector are laid out before assembly: (a) Waterproof plastic junction box 115 x 90 x 55 mm; (b) 3.7–6 V Mini diaphragm air pump; (c) DC 5V 3-Way Mini Solenoid Air Valve; (d) Mini SPST toggle switch with Pre-soldered Wires; (e) GX16 2-Pin Male Aviation Socket Connector with Wire; (f) USB-C Female Charging Jack Port; (g) 10 cm of 1/8“ ID x 3/16” OD x 1/32” Wall Ester-Based Tubing; (h) 30 cm of 1/16” ID x 1/8” OD x 1/32” Wall Ether-Based Tubing; (i) 1/16” x 1/16” x 1/16” Natural Polypropylene Tee Hose Barb 3-way Fitting; (j) 2 mm Hose Barb Bulkhead Stainless Pipe Fitting; (k) Mini PWM 4.5-35V DC Motor Speed Controller Module; (l) Twist-on quick wire connector cap; (m) USB-C 5V 3A [Raspberry Pi 4] Power Adapter; (n) Foot Pedal Switch Nonslip Momentary 2M Wire 2Pin GX16 Female Connector. B. Majority of tools required for main injector assembly: (a) Variable speed drill; (b) drill bits (top to bottom): 5/8” spade (for e), 3/8” (for f), 9/32” (for k), 15/64” (for d), 7/32” (for j), 1/8” (for pre-drilling); (c) diagonal pliers or [preferably] wire stripping tool; (d) long-nose pliers; (e) miniature screw drivers; (f) ruler; (g) Phillips head screwdriver. Optional not-shown heat gun may help soften tubing for stronger connections during assembly. C-E. Beginning of assembly. Mark (C), pre-drill (D), and drill (E) all holes in the enclosure base (7/32” and 5/8” spade holes on same end, 3/8” regular hole on opposite end) and lid (15/64” and 9/32”). Note in E, lid is inverted. F. Cut and strip all wires. G. Install toggle switch into lid, USB-C power port and GX16 plug into enclosure base. H. Cut two segments of 1/16” ID tubing to 4” (10 cm) each, one segment of 1/16” tubing to 1” (2.5 cm), and two segments of 1/8” ID tubing to 1” (2.5 cm) each. I. Connect three segments of 1/16” ID tubing to 1/16” 3-way Tee fitting, with 1” segment to the middle port. Use heat gun if available. J. Connect distant ends of 1/16” ID tubing to opposite end ports (metal and plastic) of the two solenoid valves. If needed, sand plastic port barbs to allow hose to fit over. K. Connect two segments of 1/8” ID tubing to each side port of the two solenoid valves. L. Connect outflow (middle) port of pump to solenoid with open plastic top port, and connect inflow (outboard) port to solenoid with open metal bottom port. M. Bring pump, tubing, and valve assembly into enclosure base, and connect distal end of short 1/16” ID tube segment to the 2 mm panel mount hose barb. N. Screw to install 2 mm hose barb into enclosure. O. Transfer motor casing over the top of the connected 1/16” ID tubes. P-Q. Twist connect three motor wires (red here) to the right pin wire of the GX16 socket, and install twist-on cap. R-S. Twist connect two other solenoid motor wires (black here) to one wire of the toggle switch, and install twist-on cap. T-U. Twist connect other wire of toggle switch to the ground (black wire) from USB-C port, and insert twisted wire bundle into the Negative (-) Input Power (or GND in) screw terminal of the PWM motor controller board. Secure wires by screwing terminal into locked position. V. Insert other unconnected pump motor wire (black here) to the Negative Output (-) terminal of the PWM board, and screw into locked position. W. Insert wire from left pin of GX16 socket into the Positive Output (+) terminal of the PWM board, and screw into locked position. X. Insert the positive + 5V (red) wire from the USB-C port into the Positive Input Power (or +V in) terminal of the PWM board, and screw into locked position. Y. Install PWM board onto enclosure lid, secured by its potentiometer knob shaft. Z. Close and secure lid to enclosure. Label ports, knob, and switch.
Fig 4.
Assembly of the custom home-built glass capillary needle holder.
A. All parts and additional tools required for the custom glass capillary holder: (a) 1 mm ID/ 2 mm OD PTFE tube; (b) 10 mm M4 through-hole air-out Allen socket grub/set screw; (c) 100 mm Aluminum spacer M4 round Standoff rods; (d) Mini Barb Fitting “M-4AU-3(3-M4)”; (e) M4 Allen socket wrench; (f1) Mini Barb Fitting “M-4AU-3(3-M4)”, undrilled, for 1.0 mm glass capillary; (f2) Mini Barb Fitting “M-4AU-3(3-M4)” drilled with 1/16” bit (g2) for 1.2 mm glass capillary; (f3) Mini Barb Fitting “3-M4”, undrilled, for 1.5 mm glass capillary; (f4) Mini Barb Fitting “4-M4”, drilled with 5/64” bit (g4) for 2.0 mm glass capillary; (h1/h2) Silicone O-ring CS 1 mm, OD 3 mm, for 1 or 1.2 mm glass capillary; (h3) Nitrile O-ring OD 2.5 x ID 1.5 x CS 0.5 mm, for 1.5 mm glass capillary; (h4) Nitrile O-ring OD 2.8 x ID 1.8 x CS 0.5 mm, for 2.0 mm glass capillary; (i) glass capillaries – OD 1.2 mm shown; (j) 1–2 m of 1/16” ID x 1/8” OD x 1/32” Wall Ether-Based Tubing; (k) Thread Locking adhesive for grub screw; (l) spray lubricant for drilling barbs for attaching 1.2 mm (f2/g2) and 2.0 mm (f4/g4) glass capillaries. B. Cut a small segment of PTFE tubing to 2 mm length. C. insert short segment of PTFE tubing into the open (Allen socket) end of the 10 mm M4 grub screw. D. Completely screw mini barb fitting “M-4AU-3(3-M4)” into one end of the aluminum 100 mm M4 spacer. Sparingly apply thread lock to the threads of the M4 grub screw, and begin to screw into other end of the 100 mm M4 spacer. E-F. Use Allen key to continue screwing grub screw into the aluminum M4 spacer; continue until grub screw is approximately 2 mm recessed into the M4 spacer. G. Place three of the matching-sized O-rings on one end of the glass capillary of user choice, and park them about 1 mm from the end of the capillary. H-I. Insert the matching-sized drilled or un-drilled hose barb onto the other end of the glass capillary, and bring the O-ring/capillary/barb assembly onto the open end (with recessed grub screw) of the M4 aluminum spacer. J. Screw the capillary-attached hose barb onto the end of the M4 spacer. It should tighten to stop just under 1mm from the end of the aluminum M4 spacer. If it tightens flush, remove and unscrew the grub screw less deep in the M4 spacer. If it cannot tighten securely, remove and screw in the grub screw deeper into the M4 spacer. Repeat until it tightens securely but is not completely flush with M4 spacer. K. Slightly unscrew to loosen the capillary-attached hose barb, and remove glass capillary. Whenever change in capillary size is needed, use techniques from G-K, or alternatively prepare several holders for each capillary size. L. Connect 1-2 m of 1/16” ID tubing to the opposite end of the needle holder. Connect other end of tubing to the microinjector.
Fig 5.
Experimental setup for assessment of open-source microinjector performance.
A. 1.2 mm outer diameter borosilicate glass capillaries were pulled on a micropipette puller, and manually shaped into needles without precision microforging. B. Steady-state pressure measurements in the needle holder tubing were made with needle attached, and inject held on the PWM setting shown, with either empty needle (“Air”) or needle filled with low viscosity injectate and held into lamp oil bath. C-D. With the 112.5 μm diameter needle attached, low viscosity injections were performed on “medium” PWM setting for 30 ms (C) or 60 ms (D). All micrograph linear scales show mm units.
Fig 6.
Quantitative performance of the open-source microinjector.
A. Categorical jitter plots showing all individual droplet-volume measurements across pulse durations for each condition: low-viscosity injectate (water, ~ 1 cP) at high, medium, and low pressure, and high-viscosity injectate (maple syrup, ~ 200 cP) at high pressure. Boxplots denote interquartile range and median; black dots represent individual injections. B. Continuous plots showing mean ± SE (black circles) for each pulse duration with relative standard deviation (blue circles) and maximum error (red squares) displayed on a log-scaled secondary y-axis. Linear regressions yielded R² = 0.76 (high-pressure low-viscosity), 0.94 (medium), 0.91 (low), and 0.98 (high-pressure high-viscosity). Median RSDs were ~14–16% for low-viscosity and ~9% for high-viscosity conditions. The smallest reproducible droplets measured ≈ 22–25 nL for low-viscosity/high-pressure and ≈ 70 nL for high-viscosity/high-pressure injections.
Fig 7.
Pulse Controller Configuration.
A. Block diagram of electronic control modules in the main microinjector project (below dashed line) and in the separate (optional) pulse controller (above dashed line), which facilitates tightly-timed pump operation of the microinjector to deliver controlled, reproducible injections. The pulse controller is powered together with the microinjector, at 5V via USB-C connection. Low-cost, off-the-shelf ZK-PP1K pulse generator and HW-548 P-channel MOSFET switch boards comprise the major internal components of the pulse controller. 5V supply is fed from the left footswitch pin of the microinjector via GX16 patch cable, which is high-side switched at the HW-548 MOSFET and returned to the microinjector, where it is low-side switched for motor speed modulation via PWM. The high-side switch results in timing control gated by ZK-PP1K, and the low-side switch allows controllable pressure via user-selectable dial. Two footswitches can be connected to the pulse controller to trigger 1. pulse initiation by ZK-PP1K and 2. for manual bypass operation (i.e., for aspirations or foot-controlled timing). B. Wiring diagram of the pulse controller, for user assembly. Detailed parts list is available in S3 Table. C. Beginning of assembly. In the enclosure base, drill three 5/8” spade holes for GX16 ports and one 5/8” regular holes for USB-C power port. Install three GX16 male panel mount plugs and a USB-C female power port into the enclosure base. Mark the rectangular outline of the ZK-PP1K panel mount on the enclosure lid, drill periodic (corners and every 10 mm or so) sentinel holes with the 1/8” regular bit, then connect the sentinel holes with the saw bit on the rotary tool (i.e., Dremel brand). After removing the rear panel of the ZK-PP1K module, install the module into the window you created. D. Make all connections shown in (B). D’. Expanded view of rear of ZK-PP1K module. D”-D”’. Expanded two-angle view of connections on HW-548 board.
Fig 8.
A-A’. Final assembly of the pulse controller, with exterior GX16 jacks shown. The “injector control” port is connected via GX16 patch cable to the microinjector footswitch port. “Trigger” ports (A”) are connected to footswitches, one each for 1. triggering the controlled pulses and 2. manual start/stop. B. External connections of the main microinjector and pulse controller, which should be made prior to power-on. C. Startup operation of the microinjector and pulse controller. When powered, the pulse controller begins with pulse output denoted by “OUT.” HW-548 board LED will illuminate in synchrony with the pulses as they are delivered. To terminate pulse output, press “ON” or the pulse trigger footswitch. When off (“OUT” not illuminated), the pulse output can be restarted by depressing the pulse trigger footswitch. On first start, the ZK-PP1K module may enter PWM Mode denoted by “%”, with continuous uninterrupted pulse output. For microinjector use, we desire triggered pulse mode, which can be entered by long pressing the “SET” button (>6 seconds), and is appreciated when “%” is no longer illuminated. D. In triggered pulse mode, four settings are managed on two LCD screens in ZK-PP1K that are switched with short pressing the “SET” button (>2 seconds): The “SET”-illuminated screen shows trigger delay (s) on top and pulse number (#), while the non-“SET” screen shows pulse duration (s) on top and pulse interval (s) on bottom. The + /- buttons increment or decrement the values. Typically, trigger delay is 0, pulse number is 0, and pulse duration is adjusted to desired injection duration.