The Development of a 3D Cell Culture System for Retinoblastoma
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The Development of a Three-
Dimensional Cell Culture System for
Retinoblastoma Cells
Ayling Dominguez
Glorymar Ibáñez
Dr. Hakim Djaballah
Dr. JoAnn Gensert
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Retinoblastoma
• Rare eye cancer caused by
mutations in both RB1 genes
• Tumor grows in sleeves/cuffs
– Sporadic cancer
– Hereditary cancer
Tumor
Tumor
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Workflow
• Retinoblastoma (Y79) cells incubated with NS and
without NS in a 6-well plate
Nanoshuttles
• Plate placed on magnetic dot drive to aggregate
Magnets
• Cells cultured in incubator
• Images taken to determine dimensional morphology
Culturing and Imaging
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3D Cell Culture System Assessment
Cell aggregation increases over course of study in
a ring-like formation.
Day 7Day 2 Day 3 Day 4 Day 5 Day 6Day 1
withNSwithoutNS
1000µm
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Cell Viability Assessment
2D culture: lack of uniform clumps due to lack of magnetization
3D culture: novel growth in uniform clumps, viability surrounding ring
withNSwithoutNS
Calcein AM (green channel)Brightfield
500µm
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Compound Screening
• The compounds screened were:
– 100 nM of TOPOTECAN
– 10 μM of MELPHALAN
– 100 μM of CISPLATIN
Add alamarBlue Reagent (10%
volume of culture) directly to
medium
Image plates with LEADseeker™
homogeneous imaging system
Treat cells
with compounds
Incubate
37°C
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Subsequent Research
• Screening selected compounds (topotecan,
melphalan, and cisplatin) in three-dimensional
culture system
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Acknowledgements
• Glorymar Ibáñez
• Dr. Hakim Djaballah
• Memorial Sloan Kettering Cancer Center
High-Throughput Core Screening Facility
• Dr. JoAnn Gensert