instructables

Low Cost Bioprinter

by mechbioprinter

We are an undergrad-led research team at UC Davis. We are a part of BioInnovation Group, which operates in the
TEAM Molecular Prototyping and BioInnovation Lab (Advisers Dr. Marc Facciotti, and Andrew Yao, M.S.). The lab
brings together students of diverse backgrounds to work on this project (mech/chemical/biomed engineering).

A bit of background on this project is that we started printing transgenic rice cells in collab with Dr. Karen
McDonald of the ChemE department with the goal to develop a low-cost bioprinter to make bioprinting more
accessible to research institutions. Currently, low-end bioprinters cost approximately $10,000 while high-end
bioprinters cost approximately $170,000. In contrast, our printer can be built for approximately $375.

Supplies:

Parts:

1. Ramps 1.4: https://www.amazon.com/HiLetgo-Control-Printer-Su...

2. Arduino mega 2560: https://www.amazon.com/Elegoo-EL-CB-003-ATmega2560...
3. Stepper motor drivers: https://www.amazon.com/BIQU-Compatible-Stepper-Ste...
4. Additional stepper motor (optional) https://www.amazon.com/Stepper-Motor-Bipolar-64oz-...
5. Maker beam 2 in X 1 in
6. Maker beam attachment hardware
7. M3 screws assorted sizes
8. M3 nuts x2
9. 8 mm threaded rod
10. 8 mm nut
11. 608 bearing
12. Binder clip
13. Filament
14. Monoprice V2 https://www.monoprice.com/product?p_id=15365&gcli...
15. Zip ties
16. M3 heat set nuts 2mm width

Tools:

Low Cost Bioprinter: Page 1

 1. Drill bits of various sizes
2. Hand Drill
3. Drill press
4. Hacksaw
5. Soldering iron + solder
6. Wire stripper
7. Needle nose pliers
8. Hex keys various sizes

Lab supplies:

1. Petri dishes ~70mm diameter

2. 60 ml syringe with Luer-lock tip
3. 10 ml syringe with Luer-lock tip
4. Luer-lock fittings
5. Tubing for fittings
6. T Connector for tubing
7. Centrifuge
8. Centrifuge tubes 60ml
9. Scale
10. Weigh boats
11. Autoclave
12. Beakers
13. Graduated cylinder
14. 0.1M CaCl2 solution
15. Agarose
16. Alginate
17. Methylcellulose
18. Sucrose

Software:

1. Fusion 360 or Solidworks

2. Arduino IDE
3. Repetier Host
4. Ultimaker Cura 4

Low Cost Bioprinter: Page 2

Low Cost Bioprinter: Page 3
Step 1: Selecting a 3D Printer

We chose the Monoprice MP Select Mini 3D Printer
V2 as the starting 3D printer. This printer was
selected because of its low cost and high availability.
Additionally, a highly accurate 3D model of the printer
was already available which made design easier.
This instructable will be tailored for this specific
printer but a similar process can be used to convert
other common FDM printers and CNC machines.
High accuracy model:
https://www.thingiverse.com/thing:2681912

Step 2: 3D Printing

Before disassembly of the Monoprice printer, several parts need to be 3D printed for the modification of the 3D
printer. There are versions of the paste extruders, one that requires epoxy and one that doesn't. The one that
requires epoxy is more compact but more difficult to assemble.

Low Cost Bioprinter: Page 4

Step 3: Prepare Printer for Modification

The front tower panel, bottom cover and the control panel should be removed. Once the bottom has been
removed, disconnect all electronics from the control board and remove the control board.

Step 4: Interchangeable Mount

Body 1 and Body 14 each require two heat set nuts. Body 1 is mounted to the printer frame by the two M3 bolts
hidden under the belt. The bolts can be revealed by removing the belt tensioner and pulling the belt to one side.

Low Cost Bioprinter: Page 5

 Download
https://www.instructables.com/ORIG/FDU/YP7U/K28Z642A/FDUYP7UK28Z642A.zip
…

Step 5: Z Axis Switch

The Z-axis switch is repositioned so that any length needle can be used during the homing sequence without
compensating in the software. The switch should be mounted with 2 M3 screws to the printer chassis directly
under the printhead as close to the print bed as possible.

Low Cost Bioprinter: Page 6

Step 6: Wiring

The wiring is done in accordance with the Ramps 1.4 standards. Simply follow the wiring diagram. Cut off and tin
wires as needed for the terminal blocks. Some wires may need to be extended.

Low Cost Bioprinter: Page 7

Step 7: Epoxy Extruder
While this extruder takes less time to print, it does
use epoxy which increases the total build time to over
24hrs. The 8mm threaded rod should be epoxied to
the 608 bearing and the bearing should be epoxied to
the 3D printed piece Body 21. Additionally, the nut for
the threaded rod should be epoxied to Body 40. Once
the epoxy has been fully cured, the rubber tips from
the 60ml and 10 ml syringe plungers can be fitted
over Body 9 and Body 21, respectively. An
Low Cost Bioprinter: Page 8
appropriate T fitting couldn't be found so a crude one
was made from 6mm brass tubing and solder. The
extruder acts as a hydraulic system which pushes the
Bioink out of the lower chamber of the 10 ml syringe.
Air can be evacuated out of the system by vigorously
shaking the tubes while holding the T fitting at the
highest point.
 Download
https://www.instructables.com/ORIG/FTA/L5SN/K28Z64LK/FTAL5SNK28Z64LK.zip
…

Step 8: Regular Paste Extruder

This extruder can simply be bolted together. The downside to this extruder is that it is bulkier and has high
backlash.

Low Cost Bioprinter: Page 9

 Download
https://www.instructables.com/ORIG/FUZ/4W5U/K28Z6548/FUZ4W5UK28Z6548.zip
…

Step 9: Step 9: Arduino Firmware

The Arduino needs firmware to run the stepper drivers and other electronics. We chose Marlin as it is free, easily
modified with Arduino IDE and well supported. We have modified the firmware for our specific hardware but it is
quite simple to modify for other printers because all the code is commented and clearly explained. Double click the
MonopriceV2BioprinterFirmware.ino file to open the marlin configuration files.

Download
https://www.instructables.com/ORIG/FCS/OMYB/K2WL2FVE/FCSOMYBK2WL2FVE.zip
…

Low Cost Bioprinter: Page 10

Step 10: Cura Profile

The Cura profile can be imported into Ultimaker Cura 4.0.0 and used to make high surface area meshes for use in
a profusion reactor. The generation of Gcode for the printer is still highly experimental and requires much patience.
Also attached is a test gcode for a circular profusion reactor.

Download
https://www.instructables.com/ORIG/FY5/VV0W/K28Z667C/FY5VV0WK28Z667C.zip
…

Step 11: Changing Start G-code

Paste this code into start G-code setting:

G1 Z15

G28

G1 Z20 F3000

G92 Z33.7

G90

M82

G92 E0

In Repetier, to modify start Gcode go to slicer->Configuration->G-codes->start G-codes.It is necessary to modify

the G92 Z value for each particular case. Slowly increase the value until the needle is the desired distance from
the Petri dish surface at the start of the print.

Low Cost Bioprinter: Page 11

 https://www.youtube.com/watch?v=x2ToiFLr00A

Step 12: Making the Bioink

The process for developing a Bioink suitable for an application is complex. This is the process that we followed:

Summary

The hydrogel is suitable for shear-sensitive plant cells and has open macropores to allow diffusion. The hydrogel is
made by dissolving agarose, alginate, methylcellulose, and sucrose in deionized water and adding cells. The gel is
viscous until it is cured with 0.1M calcium chloride, which makes it sturdy. The calcium chloride curing solution
cross-links with the alginate to make it sturdy. The alginate is the base of the gel, the methylcellulose homogenizes
the gel, and the agarose provides more structure since it gels at room temperature. The sucrose provides food for
the cells to continue to grow in the hydrogel.

A brief overview of some of the experiments to verify the gel

We tested different hydrogels with varying amounts of agarose and recorded its consistency, how easily it printed,
and whether it sank or floated in the curing solution. Decreasing the alginate percentage made the gel too liquidy
and it was not able to keep its shape after printing. Increasing the alginate percentage made the curing solution
work so quickly, that the gel would cure before sticking to the top layer. A hydrogel that holds its shape and doesn’t
cure too quickly was developed using 2.8 wt% alginate.

How to develop a hydrogel

Materials

Agarose (0.9 wt %)

Alginate (2.8 wt %)

Methylcellulose (3.0 wt%)

Sucrose (3.0 wt%)

Calcium Chloride .1M (147.001 g/mol)

ddH20

Low Cost Bioprinter: Page 12

cell aggregates

2 Washed & Dried Beakers

1 Mixing Spatula

Aluminum Foil

Plastic Weigh Paper

Graduated Cylinder

Procedure

Making the Hydrogel:

Low Cost Bioprinter: Page 13

 1. Measure out a specific amount of ddH20 based on how much gel solution you want to prepare. Use
the graduated cylinder to obtain a specific volume of ddH20.

2. The hydrogel solution will contain Alginate (2.8 wt %)), Agarose(0.9 wt %), sucrose (3 wt%), and
methylcellulose (3 wt%). Proper portions of the components of the hydrogel solution will be
measured using the plastic weigh paper.

3. When finished weighing out all components, add ddh20, sucrose, agarose, and lastly sodium
alginate to one of the dry beakers. Swirl to mix but do not use a spatula to mix because the powder
will stick to the spatula.

4. Once mixed, wrap the top of the beaker with aluminum foil properly and label the beaker. Add a
piece of autoclave tape to the top of the foil.

5. Put the remaining methylcellulose into the other dry beaker and wrap it in aluminum foil like the
previous beaker. Label this beaker and add a piece of autoclave tape to the top of the foil.

6. Wrap 1 spatula in aluminum foil and make sure none of it is exposed. Add autoclave tape to the
wrapped spatula.

7. Autoclave the 2 beakers and 1 spatula at 121 C for 20 minutes during the sterilize cycle. DO NOT
USE THE AUTOCLAVE IN A STERILE & DRY CYCLE.

8. Once the autoclave cycle is complete, allow the gel to cool down to room temperature and once it
has reached it, start operating in the Biological Safety Cabinet.

9. Make sure to wash your hands and arms and use proper aseptic technique once operating in the
biosafety cabinet. Also MAKE SURE to not come into direct contact with objects that will touch the
gel or be close to the gel (ex: the mixing end of the spatula, or the region of the aluminum foils that
sits over the gel)

10. In the biosafety cabinet mix the methylcellulose into the gel to get homogenous spreading. Once
done mixing, rewrap the top the mixed gel solution and place in the fridge overnight.

11. From here the gel can be used for introduction of the cells or for other uses like printing.

Adding the Cells:

Low Cost Bioprinter: Page 14

 1. Filter the cells so they’re the same size. Our procedure for filtering is

1. Lightly scrape the cells off the petri dish and use a 380 micrometer sieve to filter the
cells.

2. Gently mix the filtered cells in the hydrogel solution using a flat head spatula to avoid loss of the
mixture (that have been autoclaved).

3. After mixing the cells centrifuge out bubbles

4. From here the hydrogel is complete and can be used for printing, curing, and future experiments.

How to develop the curing solution (0.1M Calcium chloride, CaCl2)

Materials

Calcium chloride

ddH20

Sucrose (3 wt %)

Procedure (to make 1L curing solution)

1. Measure 147.01g calcium chloride, 30mL sucrose, and 1L ddH20.

2. Mix calcium chloride, sucrose, and ddH20 in a large beaker or container.

3. Submerge the gel in the curing solution for at least 10 minutes to cure.

Low Cost Bioprinter: Page 15

Step 13: Print!

In theory, Bioprinting is extremely simple; however, in practice, there are many factors that can cause failures.
With this gel, we have found that several things can be done to maximize success for our application:

1. Use small amounts of CaCl2 solution to partially cure the gel while printing,
2. Use a paper towel at the bottom of the petri dish to increase adhesion
3. Use a paper towel to evenly spread small amounts of CaCl2 over the whole print
4. use flowrate slider in Repetier to find correct flowrate

For different applications and different gels, different techniques may need to be used. Our procedure was
generated over several months. Patience is key.

Good luck if you attempt this project and feel free to ask any questions.

Low Cost Bioprinter: Page 16

Would it be possible to include a Bill of material that includes the sources? There are many sizes
of the Maker beam 2 in X 1 in, Can you clarify these specifications regarding lengths (Maker beam
& 8 mm threaded rod)? I want to ensure the 3D printed parts fit. Thanks for a posting this
instructable it is truly groundbreaking.
Why doesn't anyone ever zip up the code snippets so you jist have to download one file instead of
a bazillion?
Очень интересно у вас получилось . А как делать поджелудочную железу на этом принтере
? Или почку ? Можно напечатать ухо или нос ?
Looks like a great project, all that I seem to be lacking is a knowledge of what a bio-printer is /
does. But then what is Google for ? Keep up the good work...
Bioprinters have many applications in research and eventually the medical field. Our goal is to
create a lightweight system to produce many medicines. To achieve this, DNA would store the
information for the manufacture of the medicine. The DNA then would be used to transform rice
cells which then can be turned into a profusion reactor. This technique could eventually be used for
long term space missions as it isn’t possible to carry every medicine due to weight constraints.
This is cool as hell. Would you be able to direct a newbie to any books or sources to study if they
wanted to look into this field?
Could you include a brief bio printing for dummies in the intro? I'm highly unlikely to make this but
would like to know what sort of cells you are working with. Plant, animal, human, alien?
That’s a great idea! Hopefully we can post that soon. We currently are working to make a profusion
reactor from transgenic rice cells. The rice cells are transformed to produce the enzyme
butyrylcholinesterase. Butyrylcholinesterase is a promising antidote to Sarin which is used in
chemical warfare. Once we have further improved our printing methods we may attempt to print
cartilage.
excelente trabajo en pos de la ciencia

Nice job! Especially the Z axis zeroing with the needletip.

Thanks for the tips for the Gel-Composition.
You just did my friend's PhD project. He's very grateful.

Low Cost Bioprinter: Page 17

Great work! As a post-doctoral fellow in biology, 3D printing enthusiast and owner of an MP Select
mini 3D printer, I am very interested in this project. In which institution do you work? What projects
have you done with this setup?
We are a undergraduate run research lab at UC Davis. (https://bigucd.com/) We currently are
working to make a profusion reactor from transgenic rice cells. The rice cells are transformed to
produce the enzyme butyrylcholinesterase. Butyrylcholinesterase is a promising antidote to Sarin
which is used in chemical warfare. Once we have further improved our printing methods we may
attempt to print cartilage.

Low Cost Bioprinter: Page 18