01

Technology

The core technology of Greiner Bio-One’s Magnetic 3D Cell Culture is the magnetization of cells with biocompatible NanoShuttle™-PL. The reproducible formation of one spheroid per well in an F-bottom plate with Cell-Repellent surface is forced by magnets either by levitation or bioprinting, to form structurally and biologically representative 3D models in-vitro

Spheroid formation by magnetic levitation, bioprinting or printings of rings.

Advantages of the magnetic 3D cell culture (m3D) technology are:

  • 3D in a 2D workflow
  • Reproducible spheroid formation
  • Scalable – 6 Well to 1536 Well
  • Performed on a flat surface optimal for high-resolution microscopy and HTS
  • Rapid 3D culture formation within 24 hours for most cell types
  • No specialized equipment or media
  • Easy media changes and co-culture of different cell types
  • Compatible with fluorescence microscopy, Western blotting, qRT-PCR, Flow Cytometry, viability assays, chemiluminescence, etc.
  • Ready for automation
  • Biocompatible nanoparticles to magnetize cells

In contrast to magnetic levitation, with magnetic 3D bioprinting, the magnetized cells in a plate are printed into spheroids by placing atop a drive of magnets. One magnet below each well utilizes mild magnetic forces to induce cell aggregation and print one spheroid at the bottom of each well anywhere within 15 minutes to a few hours. Afterwards the spheroids can be cultured long-term without the use of magnetic force. This system overcomes the limitations of other platforms by enabling rapid formation of spheroids, reproducible and scalable in size for high-throughput formats (96, 384 and 1536 well) and without limitation to cell types. The 3D printing method together with commercially available standardized biochemical assay methods to facilitate continuous assessment of cell viability and other functions, provides an ideal combination for high-throughput compound screening.

By magnetized spheroids, adding and removing solutions is made easy by the use of magnetic forces to hold down spheroids during aspiration, limiting spheroid loss. Spheroids can also be picked up and transferred between vessels using magnetic tools such as the MagPen™. Magnetic forces can also be used to create co-cultures with fine spatial organization.

Bioprinted 3D co-cultures of lung adenocarcinoma (Calu-3; red) and fibroblasts (green) after 16 hours. Cancer cells reproducibly localized inside, while fibroblasts are mostly at the outside of the co-culture.
Basic steps of magnetic Bioprinting with magnetization of the cells, transfer to cell-repellent plate and bioprinting with a magnet.

Magnetic levitation is an easy tool to create native tissue environments in vitro. In magnetic levitation the magnetized cells are levitated off the bottom by a magnet above the plate. By levitating cells off the plate bottom the magnetic forces work as an invisible scaffold that rapidly and gently aggregates cells and induces cell-cell interactions and ECM synthesis. The 3D culture is formed without any artificial substrate or specialized media or equipment and can be cultured long-term. The 3D culture is formed without any artificial substrate or specialized media or equipment and can be cultured long-term.The gentle nature of magnetic levitation allows cultures to acquire macroscale morphology that mostly resembles its tissue of origin. 3D cell cultures can be analyzed using common biological research techniques, such as immunohistochemical analysis and western blotting.

The magnetic levitation has been successfully used to make 3D cultures with different cell types, including different cell lines, including stem cells and primary cells.

The current standards for compound screening are animal models; while representing human tissues of interest, these models are expensive, scarce, and carry ethical challenges. On the other end, 2D in vitro assays poorly mimic native cellular environments and thus human in vivo response, but offer high-throughput testing with ease. There is a demand for in vitro assays that are both predictive of human in vivo response and high-throughput.

As a result, we developed a viability assay, the BiO Assay. Based on magnetic 3D bioprinting, cells magnetized with NanoShuttle™-PL (NS) are printed into spheroids and rings. Immediately after printing, these structures will shrink/close, as a function of cell migration, viability, cell-cell interaction, and/or proliferation, and varies with dosage. Ring closure can be captured using a compact imaging kit (n3Dock) with an iPod™ programmed by a freely available app (Experiment Assistant) to image whole plates at specific intervals, forgoing the need to image well-by-well under a microscope. Culture contraction is generally complete within 24 hours, and images are batch processed to rapidly yield toxicity data Moreover, as the assay is label-free, the remaining rings or spheroids are available for further experimentation (IHC, Western blot, genomics, etc.).

The BiO Assay can be used to track the culture contraction of both rings and spheroids representing different situations. For rings, closure of the ring can represent wound-healing, wherein cells are working to close the void in the middle of the ring. Additionally, rings can represent similarly shaped tissues, like blood vessels, where dilation and contraction can be assayed. For Spheroids, contraction is related to spheroid assembly, with the assay macroscopically measuring how well the cells are interacting and migrating to build a competent structure.

The BiO Assay combines 3D cell culture environments with high-throughput and high-content testing to effectively predict in vivo response in vitro.

iPod-based imaging.

In contrast to standard tissue culture surfaces which are optimised to enhance conditions for cell attachment, the cell-repellent surface has been developed to effectively prevent cell attachment. CELLSTAR® cell culture vessels with a cell-repellent surface reliably prevent cell attachment in suspension cultures of semiadherent and adherent cell lines where standard hydrophobic surfaces generally used for suspension culture are insufficient.

For formation of spheroids, stem cell aggregates and self-assembled spherical clusters used as 3D cell culture models, the cell-cell interaction must dominate over the interaction between the cells and the culture surface of containment. Therefore CELLSTAR® cell culture vessels with cell-repellent surface effectively prevent cell adherence and promote the spontaneous formation of three-dimensional spheroids by gravitation: a single spheroid per well in round bottom microplates or multiple spheroids in flat bottom plates, dishes and flasks.

Long-term incubations of hydrogel cultures are frequently performed as an approach to mimic a 3D environment. When standard tissue culture vessels are used in this approach, some cells tend to migrate out of the hydrogel, forming a 2D subculture on the vessel surface. Analysis of such a cell population will therefore result in mixed data from both 2D and 3D cell cultures. CELLSTAR® cell culture vessels with a cell-repellent surface can be used for hydrogel cultures to effectively suppress the formation of 2D subcultures.

Tumor cell spheroids grown in a 96 well U-bottom CELLSTAR® cell culture microplate with cell-repellent surface.
a) LNCaP cells form single spheroids in 96 well U-bottom microplates with cell-repellent surface. 3,000 cells were seeded per well and incubated at 37° C and 5% CO2 over a 7 day period.
b) Aggregate formation of human induced pluripotent stem cells (iPSCs) cultured in a 96 well U-bottom microplate with cell-repellent surface.

02

Applications & Cell lines

What 3D platform is best for your application?

Stem Cell LinesCancer Cell LinesPrimary Cell LinesOther Cell Lines
Neural stem cellsLN229 – Glioblastoma cellsPrimary Glioblastoma cellsHuman Astrocyte
Mesenchymal stem cellHepG2 — Human liver carcinoma Human lung primary cells:
- epithelial
- endothelial
- fibroblasts
- smooth muscle		3T3 fibroblasts
Neural crest-derived mesenchymal stem cellA549 — Lung epithelial adecarcenomaValvular interstitial cells (VICs)Bend (brain endothelial)
Dental pulp stem cellPC3 — Human prostate cancerValvular endothelial cells (VECs)Adipocyte
H-4-II-E — Rat hepatoma, liverAortic valve co-cultures (AVCCs)Huvec - Human umbilical endothelial cells
MDA-231 - Human breast cancerPrimary mouse heart cellsHPF - Human pulmonary fibroblast
LNCaP - Prostate cancer cell lineHuman Primary vascular smooth muscleSMC - Tracheal smooth muscle cell
Ovarian cancer cellsPrimary Miomytrial Smooth muscleHEK293 - Human embryonic kidney
Panc-1 - Pancreatic cancer cellPrimary human hepatocytesMCF-10A - Breast epithelial cell line
Cancer associated fibroblastsPrimary pancreatic cancer cellsFibroblast
Triple negative inflammatory breast cancerPrimary tissue from PDXChondrocytes
Caki-1 – Human renal cancer cell linePrimary fibroblastsT-cells
OsteosarcomaKeratinocytesA10 - Rat vascular smooth muscle
HCT116 - colon cancer cell line

03

Products

TypeOrder no.Description
Magnetic levitation 6 Well / 24 Well6578406 Well Bio-Assembler™ Kit
Magnetic levitation 6 Well / 24 Well66284024 Well Bio-Assembler™ Kit
Magnetic levitation 6 Well / 24 Well66282424 Well Custom Lid
Spheroid Bioprinting 96 Well / 384 Well65584096 Well Bioprinting Kit
Spheroid Bioprinting 96 Well / 384 Well65584196 Well Bioprinting Kit µClear®
Spheroid Bioprinting 96 Well / 384 Well65585096 Well Ring Drive
Spheroid Bioprinting 96 Well / 384 Well781840384 Well Bioprinting Kit
Spheroid Bioprinting 96 Well / 384 Well781841384 Well Bioprinting Kit µClear®
Spheroid Bioprinting 96 Well / 384 Well781850384 Well Ring Drive
Screening & Imaging 96 Well / 384 Well65584696 Well BiO Assay™ Kit
Screening & Imaging 96 Well / 384 Well65584996 Well BiO Assay™ Kit & Imaging System
Screening & Imaging 96 Well / 384 Well781846384 Well BiO Assay™ Kit
Screening & Imaging 96 Well / 384 Well781849384 Well BiO Assay™ Kit & Imaging System
Consumables / Accessories for Magnetic 3D Cell Culture657841NanoShuttle™-PL Refill
Consumables / Accessories for Magnetic 3D Cell Culture657843NanoShuttle™-PL Refill 3 Pack
Consumables / Accessories for Magnetic 3D Cell Culture657846NanoShuttle™-PL Refill 6 Pack
Consumables / Accessories for Magnetic 3D Cell Culture657852NanoShuttle™-PL Refill 12 Pack
Consumables / Accessories for Magnetic 3D Cell Culture657850MagPen™ 3 Pack
Consumables / Accessories for Magnetic 3D Cell Culture657847NanoShuttle™-PL Refill 6 Pack with free iPod
Consumables / Accessories for Magnetic 3D Cell Culture657860Imaging Kit
Consumables / Accessories for Magnetic 3D Cell Culture657810Battery Power for Imaging Kit
TypeOrder no.Description
Dishes with Cell-Repellent Surface627979Cell Culture Dish, Ø 35x10 mm, PS, cell-repellent surface, clear, sterile
Dishes with Cell-Repellent Surface628979Cell Culture Dish, Ø 60x15 mm, PS, cell-repellent surface, clear, sterile
Dishes with Cell-Repellent Surface664970Cell Culture Dish, Ø 100x20 mm, PS, cell-repellent surface, clear, sterile
Flasks with Cell-Repellent Surface690980Cell Culture Flask, 50 ml, PS, cell-repellent surface, sterile, white screw cap
Flasks with Cell-Repellent Surface690985Cell Culture Flask, 50 ml, PS, cell-repellent surface, sterile, white filter screw cap
Flasks with Cell-Repellent Surface658980Cell Culture Flask, 250 ml, PS, cell-repellent surface, sterile, white screw cap
Flasks with Cell-Repellent Surface658985Cell Culture Flask, 250 ml, PS, cell-repellent surface, sterile, white filter screw cap
Flasks with Cell-Repellent Surface660980Cell Culture Flask, 550 ml, PS, flat flask design, cell-repellent surface, sterile, white screw cap
Flasks with Cell-Repellent Surface660985Cell Culture Flask, 550 ml, PS, flat flask design, cell-repellent surface, sterile, white filter screw cap
Flasks with Cell-Repellent Surface661980Cell Culture Flask, 650 ml, PS, high flask design, cell-repellent surface, sterile, white screw cap
Flasks with Cell-Repellent Surface661985Cell Culture Flask, 650 ml, PS, high flask design, cell-repellent surface, sterile, white filter screw cap
Multiwell Plates with Cell-Repellent Surface6579706 well Multiwell Plate, PS, cell-repellent surface, clear, with lid, sterile
Multiwell Plates with Cell-Repellent Surface66297024 well Multiwell Plate, PS, cell-repellent surface, clear, with lid, sterile
Multiwell Plates with Cell-Repellent Surface67797048 well Multiwell Plate, PS, cell-repellent surface, clear, with lid, sterile
Microplates with Cell-Repellent Surface65597096 well Microplate, PS, F-bottom/chimney well, cell-repellent surface, clear, with lid, sterile
Microplates with Cell-Repellent Surface65597696 well Microplate, PS, F-bottom/chimney well, cell-repellent surface, black, μClear®, with lid, sterile
Microplates with Cell-Repellent Surface655976-SIN96 well Microplate, PS, F-bottom/chimney well, cell-repellent surface, black, μClear®, with lid, sterile
Microplates with Cell-Repellent Surface65097096 well Microplate, PS, U-bottom, cell-repellent surface, clear, with lid, sterile
Microplates with Cell-Repellent Surface65197096 well Microplate, PS, V-bottom, cell-repellent surface, clear, with lid, sterile
Microplates with Cell-Repellent Surface65097996 well Microplate, PS, U-bottom, cell-repellent surface, clear, with lid, sterile
Microplates with Cell-Repellent Surface781970384 well Microplate, PS, cell-repellent surface, clear, with lid, sterile
Microplates with Cell-Repellent Surface781976384 well Microplate, PS, cell-repellent surface, black, μClear®, with lid, sterile
Microplates with Cell-Repellent Surface781976-SIN384 well Microplate, PS, cell-repellent surface, black, μClear®, with lid, sterile
Microplates with Cell-Repellent Surface787979384 well Microplate, PS, U-bottom, cell-repellent surface, black, μClear®, with lid, sterile

04

Publications

YearAuthor/Title/LinkReference
2018Eckhardt, B. L. et al. Clinically relevant inflammatory breast cancer patient-derived xenograft-derived ex vivo model for evaluation of tumor-specific therapies. PLOS ONE. 13, 5 (2018)
2018Hou, S. et al. Advanced Development of Primary Pancreatic Organic Tumor Models for High-Throughput Phenotypic Drug Screening.SLAS DISCOVERY. 0, 0 (2018)
2017Noel, P. et al. Preparation and Metabolic Assay of 3-dimensional Spheroid Co-cultures of Pancreatic Cancer Cells and Fibroblasts.J. Vis. Exp. 126, 56081 (2017)
2017 Desai, P. K., Tseng, H. & Souza, G. R. Assembly of hepatocyte spheroids using magnetic 3D cell culture for CYP450 inhibition/induction.Int. J. Mol. Sci. 18, 1085 (2017)
2017Souza, G. R. et al. Magnetically bioprinted human myometrial 3D cell rings as a model for uterine contractility.Int. J. Mol. Sci. 18, 683 (2017)
2016Pan, Y. et al. miR-509-3p is clinically significant and strongly attenuates cellular migration and multi-cellular spheroids in ovarian cancer. Oncotarget. 7.18, 25930-25948 (2016)
2016Tseng, H. et al. A high-throughput in vitro ring assay for vasoactivity using magnetic 3D bioprinting.Sci. Rep. 6, 30640 (2016)
2016Hogan, M. et al. Assembly of a functional 3D primary cardiac construct using magnetic levitation. AIMS Bioeng. 3, 277–288 (2016)
2016Lin, H. et al. Nanoparticle improved stem cell therapy for erectile dysfunction in a rat model of cavernous nerve injury. J. Urol. 195, 788-95 (2016)
2015Tseng, H. et al. A spheroid toxicity assay using magnetic 3D bioprinting and real-time mobile device-based imaging. Sci. Rep. 5, 13987 (2015)
2014Jaganathan, H. et al. Three-dimensional in vitro co-culture model of breast tumor using magnetic levitation.Sci. Rep. 4, 6468 (2014)
2013Tseng, H. et al. A three-dimensional co-culture model of the aortic valve using magnetic levitation. Acta Biomater. 10, 173–82 (2013)
2013Timm, D. M. et al. A high-throughput three-dimensional cell migration assay for toxicity screening with mobile device-based macroscopic image analysis. Sci. Rep. 3, 3000 (2013).
2013Haisler et al Three-dimensional cell culturing by magnetic levitationNat. Protoc. 8, 1940–9 (2013)
2013Tseng, H. et al. Assembly of a three-dimensional multitype bronchiole coculture model using magnetic levitation. Tissue Eng. Part C. Methods 19, 665–75 (2013)
2013Becker, J. L. & Souza, G. R. Using space-based investigations to inform cancer research on Earth. Nat. Rev. Cancer 13, 315–27 (2013)
2012Daquinag, A. C., Souza, G. R. & Kolonin, M. G. Adipose tissue engineering in three-dimensional levitation tissue culture system based on magnetic nanoparticles. Tissue Eng. Part C. Methods 19, 336–44 (2012)
2010Souza, G. R. et al. Three-dimensional tissue culture based on magnetic cell levitation. Nat. Nanotechnol. 5, 291–6 (2010)
YearTitleConference
2019Three-dimensional cytotoxicity assay using magnetic 3D bioprinting for measuring CAR T cell function in heterogeneous solid tumor microenvironmentsAmerican Association of Cancer Research Annual Meeting, Atlanta, GA
20183D Cultures of iPSC-derived Human Motor Neurons & Tracheal Smooth Muscle Cells in HTS Format Using Magnetic 3D BioprintingSociety for Neuroscience Annual Meeting, San Diego, CA
2018A PDX-DerivedEx-Vivo Tumor Tissue Array Platform Utilizing Magnetic 3D Bioprinting for The Identification of Tumor-Specific TherapiesAmerican Association of Cancer Research Annual Meeting, Chicago, IL
2018Validation of Magnetic 3D Spheroid Bioprinting in Combination with a BlueWasherSociety of Lab Automation and Screening Annual Conference and Exhibition, San Diego, CA
2016High-throughput spheroid formation for compound screening using magnetic 3D bioprintingDechema 3D Cell Culture, Freiburg, Germany
2016Development of spheroids derived from tumor biopsies and patient-derived xenografts using magnetic 3D bioprintingAmerican Association of Cancer Research Annual Meeting, New Orleans, LA
2016Magnetically 3D bioprinted hepatocyte spheroids for in vitro metabolic studiesSociety of Toxicology Annual Meeting, New Orleans, LA
2016High-throughput spheroid formation for compound screening using magnetic 3D bioprintingSociety of Lab Automation and Screening Annual Conference and Exhibition, San Diego, CA
2015Somatic mutation detection from liquid biopsy-derived cellular aggregates formed by magnetic 3D bioprintingAACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics, Boston, MA
2015High-throughput functional toxicity screening with iPS-cardiomyocyte and hepatocyte spheroids by magnetic 3D bioprintingCellular Dynamics iForum, Chicago, IL
2015High-throughput spheroid formation in a 384-well format using magnetic 3D bioprintingAmerican Association of Cancer Research Annual Meeting, Philadelphia, PA
2015High-throughput spheroid printing and toxicity testing using magnetic 3D bioprintingSociety of Lab Automation and Screening Annual Conference and Exhibition, Washington, DC
2014Magnetic 3D Bioprinting: A novel high-throughput and high-content assay for toxicity screeningEuropean Society of Toxicology In Vitro International Conference, Egmond aan Zee, The Netherlands
2014A novel vascular “ring” assay for smooth muscle contractility using magnetic 3D bioprintingArteriosclerosis, Thrombosis, and Vascular Biology Scientific Sessions, Toronto, ON
2014Magnetic 3D Bioprinting: A novel high-throughput and high-content assay for toxicity screeningSociety of Toxicology Annual Meeting, Phoenix, AZ
2013A high-throughput three-dimensional magnetically printed cellular assay (BiO Assay) for toxicity screening for breast cancer applicationsSan Antonio Breast Cancer Symposium, San Antonio, TX
2013A high-throughput three-dimensional cell migration assay for toxicity screening using magnetic levitation with mobile device-based macroscopic image captureAmerican Association of Pharmaceutical Scientists Annual Meeting and Exhibition, San Antonio, TX

05

Downloads

TypeOrder no.Description
SoftwareAnalysis Software for Imaging System
BrochureF0710763D Cell Culture - 3D in a fast and easy 2D workflow with products from Greiner Bio-One
Product FlyerF072100Magnetic 3D Cell Culture - Do This, Not That
Product FlyerF0731263D Cell Culture - Publications
Product FlyerF073236CELLSTAR® Cell-Repellent Surface - Cell culture vessels for suspension and spheroid culture
Product FlyerF07313796 & 384 Well Cell-Repellent U-Bottom Microplates - For high throughput 3D spheroid cell culture
One PagerF072102Advanced Development of Primary Pancreatic Organoid Tumor Models for High-Throughput Phenotypic Drug Screening
One PagerF072103Clinically relevant inflammatory breast cancer patientderived xenograft–derived ex vivo model for evaluation of tumor-specific therapies
One PagerF072104Magnetic 3D Cell Culture - Performing cell culture in 3D as easily as in 2D
ForumF073777CELLSTAR® Cell Culture Vessels with Cell-Repellent Surface
IFU -Single-Well Bio-Assembler™ Kit - REF 627840
IFU -96 Well Bioprinting Kit (clear plates) - REF 655840
IFU -96 Well Bioprinting Kit (black plates) - REF 655841
IFU -96 Well BiO Assay™ Kit - REF 655846
IFU -Battery Power for Imaging Kit - REF 657810
IFU -6 Well Bio-Assembler™ Kit - REF 657840
IFU -MagPen™ - REF 657850
IFU -Imaging Kit - REF 657860
IFU -24 Well Bio-Assembler™ Kit - REF 662840
IFU -384 Well Bioprinting Kit (clear plates) - REF 781840
IFU -384 Well Bioprinting Kit (black plates) - REF 781841
IFU -384 Well BiO Assay™ Kit - REF 781846

News / Events

Mar132019
News

3D Cell Culture at SLAS in Washington, DC

Learn about cur­rent and future appli­ca­tions in 3D cell cul­ture. At the 2019 SLAS trade show in Washington, Glauco Souza describes our 3D cell cul­ture prod­ucts and high­lights their advan­tages and fields of appli­ca­tion. Watch the video and get a deeper insight into 3D Cell Culture.

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Jan142019
News

Top 5 publication of SLAS Discovery in 2018

The pub­li­ca­tion ‘Advanced Development of Primary Pancreatic Organoid Tumor Models for High-Throughput Phenotypic Drug Screening‘ from S. Hou et al., which was cre­ated in coop­er­a­tion with the Scripps Research Institute in Florida, USA and pub­lished in SLAS Discovery in 2018, has now made it into the TOP 5 of the most read SLAS Discovery arti­cles. Please…

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Contact Persons

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Glauco Souza, Ph.D.

Director of Global Business Development & Innovation 3D Cell Culture Bioscience

Phone +1 (0)281 939-3407
[email protected]

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Dr. Jennifer Bischoff

Produktmanager / Product Manager

Phone +49 (0)7022 948-317
[email protected]

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Maybachstr. 2
D-72636 Frickenhausen
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Phone: +49 7022 948-0
E-Mail: [email protected]

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