Bioverse | Concentric 1 Cell Type

Design of Blueprint

Software used to draw 3D model

Name of software

123D Design Software was used to draw a 3D model of a square (10x10x0.8mm) and export the CAD model as an STL file. Slic3r software was used to convert the STL file to Gcode file, using the tissue model printing profile with concentric infill pattern.

Software used to convert 3D model to bioprinting protocol and toolpath

Bioinks

Biomaterial used

Company supplying biomaterial

Composition

Formulation of bioink

CELLINK Bioink by CELLINK

References

3D bioprinting of human chondrocyte-laden nanocellulose hydrogels for patient-specific auricular cartilage regeneration. Héctor Martínez Ávila, Silke Schwarz, Nicole Rotter, Paul Gatenholm. Bioprinting 2016, Volumes 1–2, 22-35.
dx.doi.org/10.1016/j.bprint.2016.08.003

3D Bioprinting Human Chondrocytes with Nanocellulose–Alginate Bioink for Cartilage Tissue Engineering Applications. Kajsa Markstedt, Athanasios Mantas, Ivan Tournier, Héctor Martínez Ávila, Daniel Hägg, and Paul Gatenholm. Biomacromolecules 2015 16 (5), 1489-1496.
dx.doi.org/10.1021/acs.biomac.5b00188

Brief description of synthesis or production process

Nanocellulose and sodium alginate-based bioink

Cells

Cell type(s)

Use your desired cell type

Cell concentration

Select a desired/relevant cell concentration for your experiments
Ccell (cells/ml) =

Calculate the amount of bioink needed for your experiments, Vbioink.
For example, the volume of bioink per construct is 100µL. If you need to print 30 constructs, then the bioink volume needed is 3mL.
Vbioink = Vconstruct × Nconstructs

Calculate the amount of cells needed.
Ncells = Ccell (1.1×Vbioink).

Resuspend the cells (Ncells) in 1/10th the volume of the bioink.
Vcell suspension = 0.1×Vbioink
For example, if Vbioink = 3mL, then Vcell suspension = 0.1×3mL = 0.3mL

Mix the cell suspension with the bioink using the CELLMIXER, following the application note, to obtain a homogenous cell distribution. Alternatively, you can use a sterile micro-spatula to mix the cells and bioink and then load the cell-laden bioink into a 3cc cartridge.
Place the cartridge (3cc), loaded with the cell-laden bioink, in the bioprinter (Printhead 1)

Culture media composition

Use the required growth and differentiation media for your cell type

Bioprinter

Model and Manufacturer

INKREDIBLE 3D Bioprinter by CELLINK

Bioprinting Technology & parameters

Pneumatic-driven microextrusion

Pressure

Nozzle type and size for PH1: Conical tip, 250µm ID (25 GA)
Printing pressure for PH1: 15-20 kPa

Printing speed

600 mm/min

Printhead temperature

Room temperature (22°C)

Printbed temperature

Room temperature (22°C)

Crosslinking process of bioink

After the bioprinting process, cover the cell-laden constructs with crosslinking solution (100mM CaCl2) and wait for 5 minutes. Remove the crosslinking solution and then rinse the constructs with culture medium. Place the constructs in multiple wellplates (eg. 24 wellplates) prefilled with culture medium (eg. 1mL per well) and incubate in standard culture conditions (37°C, 5% CO2 and 95% relative humidity). Change the media every second day or as often as needed.

Crosslinking method

UV

Bioprinting metrics

Time for bioprinting and crosslinking process per construct

35 seconds per construct

Volume of bioink per construct

0.20ml

Post-bioprinting

Time of 3D culture in vitro

Culture the 3D bioprinted cell-laden constructs for the amount of days that you wish to evaluate. If you’re interested in evaluating cell viability, a 14-day study is recommended. If you want to evaluate the extracellular matrix production or cell differentiation, we recommend at minimum a 28- or 42-day in vitro study.

Stability of constructs in vitro?

During the in vitro 3D culture, you can analyze cell viability, morphology proliferation as well as production of ECM components and gene expression.

Concentric 1 Cell Type

Description

Protocol