Name: Setting Up an Acoustic Assembly System and Cell Culture Harvesting for Generating Uniform Cell Spheroids
Uploaded: 2023-10-13T12:30:00
Description: Watch this Scientific Journal Video about Setting Up an Acoustic Assembly System and Cell Culture Harvesting for Generating Uniform Cell Spheroids at JoVE.com

setting up an acoustic assembly system and cell culture harvesting for generating uniform cell spheroids

Schaalbare generatie van uniforme celsferoïden met behulp van een 3D akoestisch assemblageapparaat

3D in vitro culture models, which provide more in vivo-like structural and morphological characteristics compared to conventional 2D culture models, have been recognized as promising systems in various biomedical applications such as tissue engineering, disease modeling, and drug screening1,2,3. As one type of 3D culture model, cell spheroids typically refer to cell aggregation, creating 3D spheroidal structures characterized by enhanced cell-cell and cell-matrix interactions4,5,6. Therefore, fabricating cell spheroids has become a powerful tool for enabling diverse biological studies.
Various techniques, including hanging drop7, non-adhesive plates8, or microwell devices9, have been developed to obtain spheroids. In principle, these methods commonly facilitate cell assembly by utilizing physical forces such as gravitational force while minimizing interactions between cells and the substrate. However, they often involve labor-intensive processes, have low productivity, and pose challenges for controlling spheroid size10,11. Importantly, the production of spheroids with the desired size and uniformity in sufficient quantity is of utmost importance to satisfy specific biological applications. In contrast to the above-mentioned methods, acoustic waves, as one type of external-force-driven technique12,13,14, have shown potential for mass manufacturing of cell spheroids with high quality and throughput, based on the principle of enhancing cell aggregation through external forces15,16,17,18. Unlike electromagnetic or magnetic forces, acoustic-based cell manipulation techniques are non-invasive and label-free, enabling spheroid formation with excellent biocompatibility19,20.
Commonly, standing surface acoustic waves (SAWs) and bulk acoustic waves (BAWs)-based devices have been developed to generate spheroids, utilizing the acoustic nodes (ANs) produced by corresponding standing acoustic fields21,22,23. Particularly, acoustic assembly devices based on BAWs, with the merits of convenient manufacture, easy operation, and excellent scalability, have gained attention for fabricating cell spheroids24,25. We have recently developed a facile BAWs-based acoustic assembly device with the ability to generate spheroids with high throughput26. The proposed device consists of a square polymethyl methacrylate (PMMA) chamber with three lead zirconate titanate (PZT) transducers arranged respectively in the X/Y/Z plane. This arrangement enables the creation of a 3D dot-array pattern of levitated acoustic nodes (LANs) for driving cell assembly. Compared to previously reported BAWs- or SAWs-based devices, which can only create a 1D or 2D array of ANs27,28,29, the present device enables a 3D dot-array of LANs for rapid cell aggregate formation within the gelatin methacryloyl (GelMA) solution. Subsequently, cell aggregates matured into spheroids with high viability within the photocured GelMA scaffolds after three days of cultivation. Finally, a large number of spheroids with uniform size were easily obtained from the GelMA scaffolds for downstream applications.

Auteur in de schijnwerpers: ontwikkeling van een steigervrije akoestische montagemethode voor hoogwaardige 3D-celsferoïde cultuur 

Driedimensionaal akoestisch assemblageapparaat voor massaproductie van celsferoïden

3D in vitro culture models, like cell spheroids, have been widely used in the fields of tissue engineering, disease modeling, and drug screening. Our research focuses on developing and invest per engineering method to fabricate spheroids or other 3D in vitro culture models with high quality for diverse biological studies. In the current protocol, the spheroids grew within the GelMA scaffold and are retrieved at the final layback, dissolving the scaffold, causing additional procedures.We will try to acoustically assemble cell aggregates in a cultural medium to enable them to grow within spheroids without scaffold. Compared to magnetic assembly techniques, acoustic waves can directly assemble cells in a label-free manner without potential toxic effects on cells. Besides, our protocol enables cell spheroids to be assembled in a three-dimensional array, substantially increasing throughput.

Watch this Scientific Journal Video about Setting Up an Acoustic Assembly System and Cell Culture Harvesting for Generating Uniform Cell Spheroids at JoVE.com

Setting Up an Acoustic Assembly System and Cell Culture Harvesting for Generating Uniform Cell Spheroids  

Het opzetten van een akoestisch assemblagesysteem en het oogsten van celkweek voor het genereren van uniforme celsferoïden

Steriliseer om te beginnen de kamer van het akoestische apparaat gedurende vijf minuten met 75% alcohol. Reinig het apparaat vervolgens met steriele PBS en bestraal het gedurende een uur met UV-licht. Monteer het steriele akoestische apparaat op een microscooptafel voor observatie van bovenaf van het inwendige van de kamer.Plaats een digitale microscoop aan de zijkant van het apparaat die vrij is van PZT-transducers, zodat het inwendige van de kamer van opzij kan worden waargenomen. Sluit de draden van de drie PZT-transducers onafhankelijk in serie aan op drie eindversterkers en drie uitgangskanalen van functiegeneratoren. Programmeer de instellingen op de functiegeneratoren voor elke PCT-transducer en specificeer parameters zoals sinusvormige golfvormen, frequentie en amplitude.Kweek 3CA-cellen in DMEM aangevuld met 10%FBS en 1%penicilline-streptomycine in een T25-celkweekkolf. Wanneer de cellen voor 80% samenvloeien, was de kweek dan twee keer met PBS. Voeg na het verwijderen van de PBS twee milliliter 0,05% trypsine-EDTA toe aan de kolf en incubeer bij 37 graden Celsius voor celloslating.Voeg twee milliliter compleet celkweekmedium toe aan de kolf om de trypsinisatie te stoppen. Breng de celsuspensie over in een buis van 15 milliliter en centrifugeer gedurende vijf minuten bij 200 G.

Research

JoVE Journal

Bioengineering

Setting Up an Acoustic Assembly System and Cell Culture Harvesting for Generating Uniform Cell Spheroids

Three-Dimensional Acoustic Assembly Device for Mass Manufacturing of Cell Spheroids

Cell spheroids are promising three-dimensional (3D) models that have gained wide applications in many biological fields. This protocol presents a method ...