Directions for Use

The DFU below is for Lifeink 200, though the exact same techniques and directions apply for the higher concentration version, Lifeink 220.

Download the full PDF version or continue reading below:

Note: Employ aseptic practices to maintain the sterility of the product throughout the preparation and handling of the collagen and other solutions.

Note: It is recommended that the collagen and other working solutions be chilled during the preparation of the collagen.

Note:  Vortexing of collagen is not recommended at any step.

Note: Ensure that NO bubbles enter the system. Bubbles in the system while mixing will turn your ink into a foam-like material.

Note: We recommend printing Lifeink^® at 2-8°C into a gelatin slurry support (FRESH method) for best results.

Cell Addition to Bioink: 
Note: Keep collagen chilled throughout these processes

1. Add the concentrated cell suspension in cell culture media to a sterile syringe.

Video for step 1

Note: We recommend a final bioink cell concentration of 5X10⁶/mL or greater.

Note: For best results, add 2 mL of cell suspension per 5 mL of collagen Bioink. Use a similar ratio for smaller volumes.

2. Place sterile coupler on the end of the syringe with the cell suspension.
3. Slowly push plunger in until media forms a slight external meniscus above the end of the coupler on the syringe.
4. Remove cap from the syringe with collagen and slowly push plunger in until collagen forms a slight external meniscus above the end of the syringe.
5. Couple the syringe with cells to the syringe with collagen. (Ensure that there are no air bubbles in the system. The “external meniscus” on both syringes helps ensure that there are no air bubbles introduced).
6. Slowly push plungers back and forth ~40 times to ensure thorough mixing. End with all of the material in the syringe to be used for printing.

Video for steps 2-6

7. The cell-laden bioink is now ready for positive displacement printers. Allow 5-10 minutes for the syringe to equilibrate back to 2-8°C before printing for best results.

Note: For pneumatic printers, transfer the collagen into an appropriate syringe using the coupler. The new syringe should have the seal inserted, but the plunger removed. Centrifuge the syringe at 2000 RPM for 1 minute after transferring the collagen to remove any air bubbles. Do not do the centrifuge step if there are cells already mixed in.

Watch this video for help

General Printing Notes:

1. To use a smaller volume of collagen, simply transfer the desired amount of collagen to another syringe, using the provided sterile coupler. To remove the air from the new syringe, you can do either of the following:
2. Centrifuge the syringe (capped) with the cap pointing up to cause the air to accumulate at the cap. Evacuate the air.
3. Centrifuge the syringe (capped) with the cap pointing down, and then use a hemostat to squeeze the syringe while pushing the plunger to allow the air to escape.

Removing air with a hemostat video

2. Complete your print within 1 hour of cell addition to syringe/bioink for best cell viability results.
3. When printing with FRESH gelatin slurry, allow the final printed structure to incubate at 37°C for 30 to 60 minutes and then replace the gelatin with media.
4. Avoid bubbles.

Product Cell Assay

Fibroblasts Day 2

Adipose Derived Stem Cells Day 1

Adipose Derived Stem Cells Day 6

Fibroblasts day 1

Fibroblasts day 5

Product Applications

Read our Bioprinting eBrochure Here

Product Bioprints

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Product References

References for Lifeink^® collagen bioinks:

Lan, X. et al. In vitro maturation and in vivo stability of bioprinted human nasal cartilage. Journal of Tissue Engineering 13, 204173142210863 (2022).

Stocco, T. D., Moreira Silva, M. C., Corat, M. A., Gonçalves Lima, G. & Lobo, A. O. Towards bioinspired meniscus-regenerative scaffolds: Engineering A novel 3D bioprinted patient-specific construct reinforced by biomimetically aligned nanofibers. International Journal of Nanomedicine Volume 17, 1111–1124 (2022).

Lee, A. et al. 3D bioprinting of collagen to rebuild components of the human heart. Science 365, 482–487 (2019).

Maxson, Eva L., et al. "In vivo remodeling of a 3D-Bioprinted tissue engineered heart valve scaffold." Bioprinting (2019): e00059.

Filardo, G. et al.Patient-specific meniscus prototype based on 3D bioprinting of human cell-laden scaffold. Bone & Joint Research 8,101–106 (2019).

Schmitt, T. Analysis and Classification of 3-D Printed Collagen-Bioglass Matrices for Cellular Growth Utilizing Artificial Neural Networks. University Thesis (2018).

Balakhovsky, Y. M., Ostrovskiy, A. Y. & Khesuani, Y. D. Emerging Business Models Toward Commercialization of Bioprinting Technology. 3D Printing and Biofabrication1–22 (2017). doi:10.1007/978-3-319-40498-1_25-1

Fox, S. et al. A simplified fabrication technique for cellularized high-collagen dermal equivalents. Biomedical Materials14,041001 (2019).

Lin, H.-H., Chao, P.-H. G., Tai, W.-C. & Chang, P.-C. 3D-printed collagen-based waveform microfibrous scaffold for periodontal ligament reconstruction. International Journal of Molecular Sciences 22, 7725 (2021).

Engberg, A., Stelzl, C., Eriksson, O., O’Callaghan, P. & Kreuger, J. An open source extrusion bioprinter based on the E3D motion system and Tool Changer to enable fresh and multimaterial bioprinting. Scientific Reports 11, (2021).

Tashman, J., et al. "In Situ Volumetric Imaging and Analysis of FRESH 3D Bioprinted Constructs Using Optical Coherence Tomography." BioRxiv. (2021) 

Product Videos

Lifeink Collagen Bioink

Bioprinted Constructs Video

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Introduction to Lifeink

Lifeink Instructional Video

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Safety and Documentation

Safety Data Sheet

Certificate of Origin

Declaration of Material Source

Product Disclaimer

This product is for R&D use only and is not intended for human or other uses. Please consult the Material Safety Data Sheet for information regarding hazards and safe handling practices.