Open-source spring-driven syringe pump with 3D-printed components for microfluidic applications

Graphical abstract


Hardware in context
Microfluidics is a technology that manipulates small quantities of fluids in the range of 10 − 9 to 10 − 18 L through channels with dimensions ranging from tens to hundreds of micrometers [1,2].Microfluidics offers advantages by allowing for analysis and experimentation with small fluid quantities, resulting in decreased expenses when utilizing costly experimental and analytical substances.Microfluidics is utilized in a variety of fields, including analyses such as electrochemical sensing [3] and environmental monitoring [4], the generation of droplets for droplet digital polymerase chain reaction (ddPCR) [5], drug screening [6,7], and cell screening [8], as well as in cell culture [9] and organ-on-a-chip system [10], DNA analysis [11,12], and fuel cells [13].Microfluidics is widely used in a variety of fields; however, the operation of a microfluidic device requires a syringe pump.
A syringe pump can cost between $350 [14] to approximately $3,500 [15] depending on the functions and specifications of the pump.The cost of conventional syringe pump can be a burden not only to labs in resources limited settings, but also labs in developed countries.To mitigate this, many efforts have been made to develop open-source syringe pumps, allowing users to build and customize syringe pumps at a minimal cost.By providing CAD file, component specs, and the code file online, users can build their own syringe pump with cost as little as $75 [16] to $400 [17].A standard form of an open-source syringe pump consists of a stepper motor, motor controller, lead screw and a frame that allows back and forth movement of a pusher block which pushes the syringe plunger of a plastic or glass syringe.One of the earlier open-source syringe pump was developed by Wijnen et al [18], demonstrating a construction of a functional pump using only open-source software, open-source 3D printer and open-source computer (Raspberry Pi).It is worth noting that this pump has been modified in different variations for different purposes.For example, a PCB for the pump controller was added to deliver a bolus of fluid across the user-specified timing for neuroscience research [19], and multiple pumps are connected to an open-source microfluidic control instrument to generate monodisperse emulsion [17].
One of the advantages of developing and using open-source syringe pumps is that modifications and upgrades using open-source computers such as Arduino and Raspberry Pi allow the pump to work as a stand-alone device that does not depend on a PC [20], imbues the pump with additional features such as wireless connectivity [21] and perform specific tasks by programming.Arduino board has been widely used for many applications such as programming the pump to deliver drugs and contrast agents for in-vivo imaging [22]; granting a precise feedback-control by integrating with a pressure sensor [23]; and being used as a programmable learning tool for developing a multichannel syringe pump [24].Furthermore, Raspberry Pi has been used to control two pumps simultaneously to generate cell lysates [25], and operating microfluidic devices for single cell sequencing [17,26].
Despite the low cost, high versatility and high expandability, constructing an open-source syringe pump can be a difficult task especially when there are many parts to buy and they have to be obtained from multiple vendors.One way to eliminate such a difficulty is to use the parts that are included in a commercial product.Baas et al. report an assembly of a programmable syringe pump using mostly the parts included in Ender3 3D printer kit.By using the parts provided in the kit, a set of three syringe pumps can be built [27].Recently, our research group developed a non-electric syringe pump that can be fabricated using the clockwork mechanism retrieved from a commercial cooking timer [28].However, the syringe pump requires specific model of commercial cooking timer to fabricate, which limits its replication.
Another way to eliminate the difficulty of gathering parts is to build a syringe pump using only a 3D printed parts.Therefore, our group recently developed a nonelectric syringe pump that can be assembled by using only 3D printed parts [29].The syringe pump was named precise, rapid-prototyped, nonelectric, torque-driven (PRNTD) pump.Unlike commercially available syringe pumps and existing open-source syringe pumps, the PRNTD pump consists only of 3D printed parts, which eliminates the need to gather commercial parts such as motors, screws, bolts and nuts that are required for the assembly.The pump operates without any electrical power, instead, it winds a flat spiral spring and is operated by clockwork mechanism to push or pull the syringe plunger for fluid delivery.Flow rate adjustment is achieved by using a specific combination of gears and syringe sizes.The PRNTD pump is, to the best of our knowledge, a first functional lab instrument that is not run by electricity and is assembled by using nothing but 3D printed parts.
Our previously published work reports the performance of the PRNTD pump and demonstrates useful and important microfluidic applications such as gradient generation and droplet generation.Although the paper also provides the DOI address from which STL files can be downloaded, additional instruction for assembling the pump can benefit its potential users.Thus, in this paper, we present a detailed step-by-step instruction on how to assemble the PRNTD pump.The fabrication cost of the syringe pump is $25-30, calculated based on the consumer price of the PLA filament used in this study.

Overview of 3D printed syringe pump
The PRNTD pump is made up of three main sections: the mainspring, the escapement, and the top layer.These sections are interconnected, with the mainspring providing torque as the pump's power source, the escapement regulating the torque from the mainspring, and the top layer converting the torque transmitted by the mainspring into linear motion by pushing the syringe plunger.
All components of the pump can be fabricated by 3D printing, which allows anyone that have basic knowledge about 3D printing to build one.After 3D printing, the pump is ready to use once all components are assembled according to the assembly instructions.The PRNTD pump comprised 27 parts and dimensions are 13.9 × 12.7 × 12.8 cm.The total time required for 3D printing all parts is 4 days, 4 h, completed over four separate runs.
Being able to build a pump using only 3D printed parts have several advantages.Firstly, users do not have to gather the parts required for assembly.Secondly, because it does not use any electronic parts, the cost is very cheap and in fact it is by far the cheapest open-source syringe pump reported [16].Additionally, the pump does not rely on electricity, enhancing its portability and accessibility for various on-site applications, whereas the majority of open-source pumps require an external power source for operation.
The spring-based pump however does have several limitations and disadvantages.The inner workings of the pump involve many freely rotating parts, which require a delicate optimization of the 3D printer used.The optimization should ensure that the gears are not tightly fitted to their posts to minimize friction.However, if the gaps between the rotating parts are too large, the gears may not mesh properly, leading to unwanted backlash.Another disadvantage of the PRNTD pump is the limited controllability of the flow rate.Unlike the open-source syringe pumps that have control panels for digital flow rate control [20,25], the PRNTD pump relies on the analog combination of gear ratio and the syringe size for setting the flow rate.This can be a daunting task when several different flow rates are required in an experiment.The analog flow rate setting also limits users from changing the flow rate in the middle of the pump's operation, which is a feature that is often used in microfluidic experiments.Thus, the PRNTD pump may not be suitable for

S.B. Park and J.H. Shin
experiments requiring the screening of many flow rates are required, but it is ideal for applications that require pumping of liquid with a specified flow rate.Lastly, because the spiral spring is 3D printed, its torque is less than that provided by a metal spring or electrically driven stepper motors.Thus, the PRNTD pump's flow rate can be affected by the back pressure, or the resistance of the system to which fluid is delivered.This results in large percent error of the flow rate, which can be as high as 27.8 % [29] However, the accuracy ideally should be in a single-digit value, preferably within ± 5 % [18].In future research, PRNTD pump can be redesigned to use a threaded screw, limiting the amount of torque delivered to the spring from the back pressure, thereby improving accuracy.

PRNTD pump
• Researchers can build the syringe pump to operate the microfluidic devices that require consistent fluid flow.
• Researchers can build the syringe pump only with 3D printing and fabricate it for $25-30.
• The syringe pump operates without electricity using a 3D printed spiral spring, eliminating the need for additional components like motors and microprocessors.• The syringe pump is lightweight, portable and can be used in low-resource settings with limited power supplies.

Table 1
Label, part name, and description for the components of the mainspring section.

Design files
Mainspring section.The Mainspring section corresponds to the power supply of a commercial syringe pump.In the PRNTD pump, unlike a commercial syringe pump that depend on electricity to run a motor, the torque from the mainspring acts as the driving force.By rotating the handle of the PRNTD pump clockwise, the mainspring is wound up, and the torque generated as the mainspring unwinds serves as the power source for the syringe pump.The Mainspring section consists of the mainspring, spring barrel holder, spring barrel bottom, and spring barrel top (Table 1).
Escapement section.The Escapement section operates by utilizing the torque generated as the mainspring unwinds.This torque, transmitted through the torque reduction gear, rotates the escapement wheel, causing the pallet fork to oscillate.The pallet fork, in turn, imparts a pendulum-like motion to the balance wheel, ensuring a consistent unwinding speed of the mainspring.The Escapement section is comprised of the escapement frame, main gear, ratchet, torque reduction gear 1 and 2, escapement wheel, hairspring, pallet fork, and balance wheel (Table 2).
Top layer section.The top layer plays a role in adjusting the forward or backward speed of the pusher block in a commercial syringe pump.The torque from the mainspring is transmitted to the rack gear through the arrangement of gears, converting it into linear motion.The forward speed of the rack gear is adjusted to push the syringe plunger by controlling the gear ratio.The forward speed of the rack gear can be controlled by changing the gear ratio, accomplished by combining a total of four gears.In the syringe pump, different-sized syringes can be assembled by mounting a syringe holder.This includes 1-, 3-, or 5-mL disposable syringes, as well as 50-, 100-, 250-, or 500 µL Hamilton gastight glass syringes.Depending on the desired flow rate, different sizes of syringes can be mounted.The top layer section consists of a top layer frame, two main gear holding frames, four gear posts, a syringe holder, and a winding key in both forward pumping mode and reverse pumping mode.It is important to note that a forward pumping mode requires four velocity reduction gears, whereas a reverse pumping mode requires five velocity reduction gears (Table 3).To change the forward speed of the syringe plunger for flow rate adjustment, user can alter the positions of the gear posts and then assemble the velocity reduction gears of different numbers of gear teeth.Lastly, one of the six rack gears is selected according to the target gear ratio and syringe type.A disposable syringe requires mounting of rack gear R4-D1, R4-D2, or R4-D3 and syringe holder R6-P, whereas a glass syringe requires mounting of rack gear R4-G1, R4-G2, or R4-G3 and syringe holder R6-G.The detailed instructions explaining the correct combination of the velocity reduction gears and rack gears is shown in Table 7.

Bill of materials summary
For the fabrication of all components of the syringe pump, it requires 45.28 m (358 g) of PLA, including support for the printed objects.The total cost of building a syringe pump is $25-30 (Table 5).

Printing settings
The quality and characteristics of 3D-printed objects, such as the PRNTD pump, depend on factors like the resolution of the 3D printer, material properties used in 3D printing, the minimum achievable feature size determined by slicing conditions, and the accuracy and resolution of the 3D-printed parts.Table 6 shows the 3D printer setting values used by the authors for the fabrication of the    PRNTD pump.These 3D printing settings are also presented at supplementary files of previously published research [29].The software Ultimaker Cura slicer (Ultimaker, Netherlands) was used for slicing and setting the print.Ultimaker S3 (Ultimaker, Netherlands) was used to build PRNTD pump in this study.

Mainspring assembly
1) Assemble spring barrel bottom (M2) onto the spring barrel holder (M1) in a cross-shaped configuration (Fig. 1A).Align the post on M2 (Fig. 1B) with the hole next to 'S' marked on M1 (Fig. 1C).2) Slightly wind the mainspring (M3) to reduce its outer diameter, and while preventing the mainspring from unwinding, place M3 into M2 by assembling the looped end of the M3 with one of the prongs of M1.Make sure to insert the prong that is marked with 'S' into the looped end of M1 as shown in Fig. 1D.3) Position spring barrel top (M4) by aligning the 'S' marked hole with the prong marked 'S', then finish the assembly as shown in Fig. 1E.

Escapement assembly
1) Assemble balance wheel (E7) and hairspring (E8) (Fig. 2A and Fig. 2B).Assemble the E7 and E8 according to Fig. 2C, ensuring that the rectangular post of E7 fits into the rectangular gap of E8. 2) Assemble two torque reduction gears (E4-1 and E4-2), and escapement wheel (E5) onto escapement frame (E1) in sequence (Fig. 3A, Fig. 3B and Fig. 3C).3) Assemble the assembled E7 and E8 onto E1 (Fig. 3D).Be cautious since E1′s posts are thin; assemble the hole at the end of E8′s strip onto E1′s post (P1) marked on Fig. 3D cautiously.4) Position pallet fork (E6) with the surface labeled 'U' facing upwards.Assemble E6 onto E1 according to Fig. 3E.Ensure that E6 and banking pin of E7 engage together.5) Assemble the assembled mainspring section onto the longest post of E1 (Fig. 3F).Ensure that the post marked 'S' aligns between the three posts on E1.Make sure that the elevation of the smaller gear of E4-1 is above the assembled complex of M1-M4.6) Assemble ratchet (E3) onto the longest post of E1 (Fig. 3G).Adjust the direction of E3 to fit into the rectangular gap of M1.

7)
Assemble main gear (E2) onto E3, make sure that E3′s gear fits inside the internal gear of E2.Also ensure that E2 and the small gear of E4-1 mesh together (Fig. 3H).

Top layer section assembly
1) Assemble top layer section frame (R1) onto E1.Verify the positions of E1′s posts and the holes in R1 before assembly (Fig. 4A).

Forward pumping
The flow rate varies depending on the forward speed of the rack gear.Therefore, to adjust the forward speed, different gear ratios must be applied.Changing the gear ratio involves combining gears with different numbers of teeth, resulting in variations in the assembly positions of gears and gear posts.The Table 7 informs the assembly positions of gear posts and the corresponding gears to be assembled based on the gear ratio.By referring to Table 7, assemble R3-1, R3-2, R3-3, and R3-4 in their respective positions 1-4 (marked in the scheme as circled numbers).

Reverse pumping
In this study, to demonstrate a reverse pumping, a gear post was fabricated to enable reverse pumping with the gear ratio is 1:81 as an example.Since Reverse pumping adds a gear that changes the direction of the rack gear, distance from the last gear to the position where the rack gear is fixed should be decreased.Therefore, to achieve different gear ratios, new rack gears need to be designed  considering the distance from the last gear to the position where the rack gear is fixed.The assembly process is identical to the forward pumping, except for the substitution of R3-5 for R3-4 and the additional assembly of R5 which is a gear with 12 gear teeth.Fig. 5. illustrates the assembly positions of gear posts and the corresponding gears to be assembled based on the gear ratio.

Syringe assembly
1) Assemble winding key (R7) onto E3 (Fig. 6A) 2) Assemble the syringe plunger onto R4 and syringe barrel onto R6.In this process, assemble the syringe barrel into the gap of R6 based on the length of the syringe plunger (Fig. 6B and 6C).

Final assembly
Fig. 7 shows the assembled model of the PRNTD pump.Video S1 shows the entire assembly procedure of the PRNTD pump.

Operation instructions
Syringe pump operation instructions 1) Fix E7 (balance wheel) to prevent movement of the balance wheel while winding up the mainspring.(Placing a small weight, for example, a pen on top effectively hinders its movement).2) Rotate the R7 clockwise until M1 is fully wound.
3) To minimize backlash (the gap between the gears that needs to be closed in order for the track gear to begin pushing the syringe), remove R5-4 and rotate R5-3 counterclockwise (for forward pumping) to allow fluid to be dispensed from the syringe.When the dispensing of the fluid is observed, hold on to R5-3 while reassembling R5-4.4) Release the fixation on E7 to allow the operation of the escapement.User may need to give a nudge to E7 to initiate movement.5) For preconditioning of a newly printed spring, it is recommended to go through two cycles of operation (two repeats of winding and unwinding of the spring in an assembled pump) [27].6) A spring is recommended for a maximum of 30 cycles of use [27].

Validation and characterization
To characterize the syringe pump, we measured the flow rate.To obtain the flow rate, the dispensed weight from the syringe was measured by an electronic balance (PIONEER®, Ohaus, New Jersey, USA), and then the flow rate was calculated based on the dispensed water over time after converting weight to volume.The characterized flow rate of PRNTD pump is reported in previous work [29].

Characterization of the flow rate
The flow rate is the product of the cross-sectional area of the internal diameter of the syringe barrel and the velocity at which the syringe pump moves the syringe plunger.Therefore, the velocity of the syringe plunger and the size of the syringe affect the flow rate.The variation in flow rate based on syringe size and the velocity of the syringe plunger can be found in the original research paper.In this paper, flow rate of forward pumping and reverse pumping were characterized using different syringe sizes.Experiment was performed three times for each condition.Fig. 8A presents the dispensed volume over time measured using a 5 mL syringe.The theoretical flow rate under these conditions is 16.00 µL/min.The flow rates for forward pumping and reverse pumping were 12.85 ± 0.32 µL/min and 10.50 ± 1.57 µL/min, respectively.The R 2 values obtained from the linear fitting were 0.9841 and 0.9569.Fig. 8B shows the dispensed volume over time using a 3 mL syringe, with a theoretical flow rate of 9.3 µL/min under these conditions.The flow rates for forward pumping and reverse pumping were 6.56 ± 0.54 µL/min and 6.27 ± 1.04 µL/min, respectively, with linear fitting R 2 values of 0.9340 and 0.9982.The R 2 value indicates how close the data is to a linear regression graph, and the closer the R 2 value is to 1, the more linear the data graph is.Therefore, if the R 2 value is close to 1, the increment of dispensed volume per time is linear and consistent, which indicates the pumping is maintained without decreasing over time.
S.B. Park and J.H. Shin on next page) S.B. Park and J.H. Shin

Fig. 2 .
Fig. 2. (A), (B) Showing the exploded view of the balance wheel and hairspring assembly.(C) Schematic showing the upside and downside of the assembled balance wheel and hairspring.

Fig. 4 .
Fig. 4. Schematics showing the top layer section assembly orders of forward pumping.

Fig. 5 .
Fig. 5. Schematics showing the top layer section assembly orders of reverse pumping.

Fig. 6 .
Fig. 6.Schematics showing the top layer section assembly orders of reverse pumping.

Table 2
Label, part name, and description for the components of the escapement section.
(continued on next page) S.B. Park and J.H. Shin

Table 3
Label, part name, and description for the components of the top layer section.

Table 4
Design files summary.

Table 5
Bill of materials summary.

Table 6
3D printing setting of the syringe pump.

Table 7
(continued ) Gear ratio Scheme showing the insert locations of velocity reduction gear post and velocity reduction gear Corresponding position number of velocity reduction gears and rack gear