Gibco™ OncoPro™ Tumoroid Culture Medium Kit

Catalog No.	Quantity
A5701201	1 Kit

Catalog No. A5701201 Supplier Gibco™ Supplier No. A5701201

Description

The OncoPro Tumoroid Culture Medium Kit is an easy-to-use culture system optimized for the derivation and expansion of patient-derived tumoroid lines.

The Gibco OncoPro Tumoroid Culture Medium Kit is an easy-to-use culture system optimized for the expansion of patient-derived tumoroid lines. Tumoroids (also referred to as cancer organoids or tumor organoids) are known to have complex culture requirements that can be challenging. OncoPro medium simplifies tumoroid culture to make physiologically relevant cancer models more accessible.

Features include:

• Optimized formulation for patient-derived tumoroid culture
• Streamlined media preparation, culture methods, and support
• Adaptable use for multiple cancer indications and downstream assays

Optimized performance

OncoPro Tumoroid Culture Medium is a novel, serum-free media system specifically developed for patient-derived cancer cell expansion. Over 40 conditions were tested during its development to identify the combination of basal medium, supplements, and growth factors that would best support long-term maintenance of donor-specific characteristics, including mutational profiles and gene expression patterns. The absence of Wnt-3A, R-spondin, Noggin, and small molecule inhibitors make OncoPro medium well-suited for complex biological studies and screening applications. Meanwhile, our strict ISO 13485 and cGMP quality standards help ensure that this ready-to-use media system provides consistency and reproducibility.

Streamlined adoption

This versatile media system includes a four-component kit that can be adapted for different cancer indications through addition of tissue-specific growth factors. An innovative suspension culture method eliminates the requirement for manual cell embedding in basement membrane extract domes, helping provide a cost-effective and less labor-intensive option for routine tumoroid culture. OncoPro medium is broadly compatible with various starting materials, including dissociated tumor cells (DTCs), patient-derived xenograft organoids (PDXOs), and tumoroid lines derived in other media systems. Comprehensive resources and support are available to help ensure effortless adoption.

Adaptable application

The OncoPro Tumoroid Culture Medium Kit is a flexible system designed to help address complex research needs for long-term growth of tumoroid cell lines, and accommodates multiple cancer indications (colorectal, lung, pancreatic, head and neck, endometrial, breast) using a single media kit. Other epithelial cancer indications may also be supported. The kit’s scalable and automation-friendly suspension workflow facilitates smooth transitions to downstream research applications, including traditional 2D assays, diverse characterization techniques, and advanced cell-based modeling.

Specifications

• Cell Type: Cancer, Tumoroids, Cancer Organoids
• Classification: Serum-free, Animal Origin, cGMP, Human Origin
• Form: Liquid
• Serum Level: Serum-free
• Sterility: Sterile-filtered
• Sterilization Method: Sterile-filtered
• With Additives: Bovine Serum Albumin (BSA)
• Quantity: 1 Kit
• Culture Type: Mammalian Cell Culture, Suspension Cell Culture, 3D Cell Culture

Frequently Asked Questions (FAQs)

Can I use OncoPro Tumoroid Culture Medium Kit to derive new tumoroid lines from patient-derived tissue such as tumor resections?

OncoPro Tumoroid Culture Medium Kit has been developed and optimized for the expansion and maintenance of existing tumoroid cultures. It has been used to successfully derive new tumoroid lines from patient tumor resections, dissociated tumor cells, and patient-derived xenografts, however the success rate of new derivations is highly variable and dependent on the quality of the starting tissue, tissue type, donor, etc.

I just prepared the complete OncoPro Tumoroid Culture Medium and it appears yellow. Is this normal?

The complete OncoPro Tumoroid Culture Medium may appear yellow when it is first prepared. This is normal and does not indicate any issues with buffering or pH. The yellow color will dissipate somewhat after refrigeration and/or incubation.

When using OncoPro Tumoroid Culture Medium, do I need to add ROCK Inhibitor Y-27632 at every feed?

Yes, the complete OncoPro Tumoroid Culture Medium should be supplemented with freshly thawed 10 µM Y-27632 for every feed.

What dissociation reagent do you recommend for passaging tumoroids?

We recommend using StemPro Accutase Cell Dissociation Reagent (Cat. No. A1110501) for passaging tumoroids.

What is the recommended passaging frequency for tumoroids? At what size are tumoroids big enough to passage?

Tumoroid cultures should be passaged when the average tumoroid size is 200-300 µm in diameter. This is typically every 7 days, but depending on the tumoroid line, may occur anywhere between 4-14 days after seeding. We do not recommend passaging when the average tumoroid size is <100 µm or >400 µm in diameter.

Can I passage tumoroids early if I need to start an experiment sooner?

We do not recommend passaging tumoroids before they reach at least 200 µm in diameter. Over-dissociation can negatively impact the viability and subsequent expansion of tumoroid cultures.

How do I determine the correct split ratio when culturing tumoroids? What is the recommended seeding density? Is there a difference in seeding density for embedded and suspension cultures?

For colorectal cancer tumoroid lines, we recommend a split ratio of 1:7. We recommend a seeding density of 0.125 x 10E6 cells/mL to 0.2 x 10E6 cells/mL for suspension culture. We recommend a seeding density of 5 x 10E4 cells/dome for embedded cultures for 50µL domes.

Why do you recommend culturing tumoroids in suspension conditions?

Suspension conditions eliminate the need for manually embedding cells in basement membrane extract domes. As a result, suspension cultures are less labor intensive, more cost efficient, more scalable, and more compatible with automation than embedded cultures.

We have not found any significant differences in morphology, growth rate, mutational profile, or gene expression signature between tumoroids cultured in parallel, in suspension and embedded culture conditions.

Can tumoroids be frozen? How should I thaw tumoroids?

We routinely freeze tumoroids at 1-2 x 10E6 cells/mL in Recovery Cell Culture Freezing Medium. Please see the user manual (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0029372-OncoPro-Tumoroid-Culture-Medium-UG.pdf) for guidance on recovering cryopreserved tumoroids.

How stable are tumoroids? How long can tumoroid cultures be maintained?

We have data demonstrating that morphology, growth rate, mutational profile, and gene expression profile are conserved in tumoroid lines cultured for over 250 days (35+ passages post-thaw from bank).

What type of plates should I use for suspension or embedded cultures with OncoPro Tumoroid Culture Medium Kit?

For suspension culture, we recommend the use of Nunc Non-Treated Multidishes and Nunc Non-treated Flasks, specifically Nunc Non-Treated 6-well plate (Cat. No. 150239), Nunc Non-treated T25 EasyFlask, Filter Cap (Cat. No. 169900), and Nunc Non-treated T75 EasyFlask, Filter Cap (Cat. No. 156800). For embedded culture, we recommend the use of Nunc Cell-Culture Treated Multidishes, specifically Nunc Cell-Culture Treated 6-well plate (Cat. No. 140675).

Can OncoPro Tumoroid Culture Medium Kit be used to culture healthy tissues?

OncoPro Tumoroid Culture Medium Kit has been developed and optimized for the culture of primary patient-derived tumoroids. It has not been validated for the culture of healthy primary tissues or cells.

Does OncoPro Tumoroid Culture Medium include a Wnt activator?

OncoPro Tumoroid Culture Medium does not contain any activators of the Wnt signaling pathway.

What growth factors are present in OncoPro Tumoroid Culture Medium?

OncoPro Tumoroid Culture Medium Kit is a novel media system that was specifically developed for the expansion of patient-derived tumoroids. The exact formulation of OncoPro Tumoroid Culture Medium is proprietary, however unlike other published methods, OncoPro Tumoroid Culture Medium does not rely on the addition of Wnt, R-spondin, or Noggin for long term maintenance of patient-derived tumoroids. OncoPro Tumoroid Culture Medium also does not contain any small molecule inhibitors.

Can I use OncoPro Tumoroid Culture Medium Kit for multiple different cancer indications?

Yes, OncoPro Tumoroid Culture Medium Kit has been validated for the culture of colorectal, lung, pancreatic, and head and neck tumoroids. Some cancer indications may require supplementation with additional growth factors.

What additional growth factors are needed to grow different cancer indications using OncoPro Tumoroid Culture Medium Kit?

The requirements for additional growth factors with OncoPro Tumoroid Culture Medium depend upon the type of cancer. Colorectal tumoroids do not require supplementation with any additional growth factors. Lung and head and neck tumoroids require the addition of heat stable FGF10. Pancreatic tumoroids require the addition of heat stable FGF10 and gastrin. The requirements for other cancer indications will need to be determined empirically.

Can I make and freeze aliquots of the components of the OncoPro Tumoroid Culture Medium Kit or complete OncoPro Tumoroid Culture Medium and thaw as needed?

Yes, we recommend preparing aliquots of the OncoPro Supplement and B-27 Supplement and storing at -20 degrees C. Please see the user manual (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0029372-OncoPro-Tumoroid-Culture-Medium-UG.pdf) for guidance on aliquoting and freezing. Avoid additional freeze-thaw cycles. We do not recommend preparing and freezing complete OncoPro Tumoroid Culture Medium.

What is the shelf life of the complete OncoPro Tumoroid Culture Medium?

The shelf life of the complete OncoPro Tumoroid Culture Medium is 1 week at 4 degrees C.

How can I transition from a competitor media system into OncoPro Tumoroid Culture Medium suspension culture?

Some tumoroid lines can be immediately transitioned into OncoPro Tumoroid Culture Medium suspension culture. Some more sensitive lines may benefit from a more gradual, stepwise transition. You can find additional guidance in the OncoPro Tumoroid Culture Medium Kit user manual (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0029372-OncoPro-Tumoroid-Culture-Medium-UG.pdf).

Is OncoPro Tumoroid Culture Medium Kit compatible with tumoroids that have been grown in other media systems?

Yes, we have successfully transitioned tumoroids that were previously cultured in a number of different media systems into OncoPro Tumoroid Culture Medium.

What is the feeding schedule for tumoroids?

We recommend feeding cells every 2-3 days until tumoroids reach an average size of 200-300 µm, at which point they are ready to be passaged or harvested for downstream assays.

Can I use wild type FGF10 instead of heat stable FGF10 for culturing my lung/head and neck/pancreatic tumoroids?

We recommend using heat stable FGF10 for cancer indications requiring supplementation with FGF10. This allows for the use of lower concentrations of growth factor and is more cost effective than using wild type FGF10. If wild type FGF10 is used, concentrations should be increased to 100 ng/mL throughout the protocol. We have successfully cultured tumoroids in medium containing 100 ng/mL wild type FGF10 without causing significant differences in morphology, growth rate, mutational profile, or gene expression signature compared to culturing in medium with 10 ng/mL heat stable FGF10.

Which extracellular matrix (ECM) should I use for culturing tumoroids in suspension?

We have successfully used various basement membrane extracts (BMEs) for culturing tumoroids in suspension. BME used in these workflows must have a concentration >10 mg/mL prior to dilution. Some ready-to-use formulations may not be suitable for these applications due to their low protein concentration.

Do tumoroids cultured in suspension culture still require the addition of basement membrane extract (BME)?

Yes, the protocol calls for the addition of 2% v/v diluted basement membrane extract (BME) to suspension cultures. This promotes the formation and stability of 3D tumoroids. Tumoroids are still free-floating at this concentration as it is much lower than the concentration used for embedded cultures.

Will transitioning of my embedded tumoroids to suspension culture conditions change my cells?

We have data demonstrating that morphology, growth rate, mutational profile, and gene expression profile are conserved between cells grown in embedded and suspension culture conditions.

Will my suspension-cultured tumoroids form xenografts in mice?

Yes, we have data showing that some tumoroids cultured using OncoPro Tumoroid Culture Medium Kit are tumorigenic and can generate tumors in mouse xenograft models. Xenograft success rates will depend on the characteristics of the tumoroid model and the mouse strain used.

Can I transition my suspension-cultured tumoroids back to 2D culture, e.g., for 2D invasion-migration assays?

Yes, we have supporting protocols available for transitioning suspension tumoroid cultures to traditional 2D assays such as migration-invasion assays.

For Research Use Only. Not for use in diagnostic procedures.