This video demonstrates the procedure of extracellular matrix-derived microcarrier fabrication by electrospraying. These microcarriers simulate the native tissue-specific microenvironment and are stable in long-term 3D in vitro cell culture models.
Protocol
1. ECM-derived Microcarrier Fabrication via Electrospraying NOTE: An overview of the electrospraying set-up is shown in Figure 1. Incubate a previously prepared cryomilled ECM suspension at 37 °C with continuous agitation at 100 rpm O/N. NOTE: Cryomilled ECM is used to fabricate the ECM-derived microcarriers to ensure greater uniformity in t…
Representative Results
Figure 1. Overview of the electrospraying apparatus used in microcarrier fabrication. A: Image showing the arrangement of the key electrospraying equipment including the syringe pump and high voltage power supply, as well as the positioning of the needle relative to the Dewar of liquid nitrogen. B: Electrospraying schematic, including the recommended ranges …
Divulgaciones
The authors have nothing to disclose.
Materials
Alligator clip leads
VWR
470149-728
Aluminium foil
Fisher Scientific
01-213-101
Centrifuge tubes (15 mL)
Sarstedt
62.554.205
Centrifuge tubes (50 mL)
Sarstedt
62.547.205
Dessicator
Fisher Scientific
8624426
For lyophilized ECM and bioscaffold storage
Dewar flask
Fisher Scientific
10-196-6
Low form; volume range of 250 – 500 mL
ECM (Extracellular Matrix)
Isolated from human adipose tissue, porcine dermis or porcine myocardium, as described in Flynn et al. 2010, Reing et al. 2010, and Wainwright et al. 2010 (ref # 28, 8, 32)
Liquid nitrogen
For electrospraying
Lyophilizer
Labconco
7750021
FreeZone4.5
Orbital incubator shaker
SciLogex
8.3201E+11
Temperature controlled (37 °C)
Retort stand
VWR
470019-526
Retort stand clamp
VWR
21573-606
Safety-Lok Syringe
BD
309606
3 mL Luer lock syringe for microcarrier fabrication and dispensing ECM suspension
Scoopula
VWR
89259-968
For collecting microcarriers
Syringe pump
VWR
10117-490
Microprocessor controlled
High voltage power supply
Gamma High Voltage Research
ES30P-5W/DDPM
Capable of recommended 15 – 25 kV voltage range
Winged infusion set
Terumo
22258092
30 cm tubing length, 25 G 3/4" recommended; Other needle gauges can be used and may influence microcarrier diameter
Bioscaffold Fabrication by Electrospraying: A Technique to Generate Microcarrier Scaffolds Derived from Extracellular Matrix of Decellularized Tissue. J. Vis. Exp. (Pending Publication), e20486, doi: (2023).