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Related Experiment Video

Updated: Apr 3, 2026

Mammary Epithelial and Endothelial Cell Spheroids as a Potential Functional In vitro Model for Breast Cancer Research
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A Simple and Cost-Effective Method for Generating Spheroids From Triple-Negative Breast Cancer Cell Line

Ramón Cervantes-Rivera1,2, Luisa Nirvana González-Fernández1, Atalia Ziret Romero Rosas1

  • 1Centro Multidisciplinario de Estudios en Biotecnología (CMEB), Facultad de Medicina Veterinaria y Zootecnia, Universidad Michoacana de San Nicolás de Hidalgo. C.P. 58893, Morelia, Michoacán, México.

Bio-Protocol
|April 2, 2026
PubMed
Summary
This summary is machine-generated.

Researchers developed a simple, cost-effective 3D spheroid model for breast cancer research. This accessible 3D tumor microenvironment model aids drug screening and understanding aggressive triple-negative breast cancer.

Keywords:
3D cell cultureDrug screeningSpheroidsTriple-negative breast cancerTumor microenvironment

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Area of Science:

  • Oncology
  • Biotechnology
  • Cell Biology

Background:

  • Breast cancer (BC) is a leading cause of cancer mortality globally.
  • Triple-negative breast cancer (TNBC) lacks targeted therapies due to its aggressive nature.
  • Current 2D cell cultures do not fully replicate the 3D tumor microenvironment (TME).

Purpose of the Study:

  • To develop a robust, simple, and cost-effective protocol for generating uniform 3D breast cancer spheroids.
  • To create a more physiologically relevant preclinical model for studying tumor biology and drug response.
  • To bridge the gap between 2D cultures and in vivo studies for improved therapeutic discovery.

Main Methods:

  • Utilized a lab-made agarose coating in 96-well plates to create non-adherent surfaces.
  • Employed a brief orbital shaking step to promote spheroid formation from MDA-MB-231 TNBC cells.
  • Implemented quantitative, image-based size analysis for monitoring spheroid growth and ensuring reproducibility.

Main Results:

  • Successfully generated uniform 3D spheroids within 96 hours using basic cell culture equipment.
  • Demonstrated the protocol's reproducibility and scalability for diverse cell types.
  • Established a clear workflow for spheroid formation to quantitative size analysis using ImageJ and GraphPad Prism.

Conclusions:

  • The developed 3D spheroid protocol provides an accessible and reliable model of the TME.
  • This model facilitates high-throughput drug screening and mechanistic studies in breast cancer.
  • The protocol supports the advancement of personalized medicine strategies for breast cancer and other malignancies.