Tissue Models

2 Cell Types

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Description

Protocol

Design of Blueprint

Software used to draw 3D model

Name of software

BioCAD

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

Spacing between lines

Line space = 2mm

Bioinks

Biomaterial used


Company supplying biomaterial


Composition

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

Cells

Cell type(s)

Primary human dermal fibroblasts (HDF) from ATCC

Primary human epidermal keratinocytes (HEK) from Life Technologies

Cell concentration

First two layers with HDF: 5x106 cells/ml. A suspension of 16.5x106 HDFs in a volume of 300µL were mixed with 3mL of CELLINK bioink using the CELLMIXER to obtain a final concentration of 5x106 cells/ml.

Last two layers with HEK: 5x106 cells/ml. A suspension of 16.5x106 HEKs in a volume of 300µL were mixed with 3mL of CELLINK bioink using the CELLMIXER to obtain a final concentration of 5x106 cells/ml.

Culture media composition

HDF growth medium: DMEM with GlutaMAX supplemented with 10% fetal bovine serum (Biochrom) and 1% penicillin–streptomycin

HEK growth medium: EpiLife® Medium supplemented with human keratinocyte grown supplement (Life Technologies)

3D cell culture medium: HDF and HEK growth medium (1:1 ratio)

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
Nozzle type and size for PH2: Conical tip, 250µm ID (25 GA)
Printing pressure for PH2: 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, the cell-laden skin constructs were crosslinked by submerging in an ionic solution of 100mM CaCl2 for 5 minutes. The constructs were then rinsed and incubated in 3D cell culture medium in standard culture conditions (37°C, 5% CO2 and 95% relative humidity).

Crosslinking method

UV

Bioprinting metrics

Time for bioprinting and crosslinking process per construct

34 seconds per construct

Volume of bioink per construct

0.20mL

Post-bioprinting

Time of 3D culture in vitro

14 days

Results

Notes

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