Liver cancer spheroid stained using CellO-IF technology.
CellO-IF: The Smarter Way To Label
All-in-one Reagent for Immunofluorescent Labelling of Organoids, Spheroids and Cells
Simplify the steps of immunofluorescence labelling and maximize your results. The CellO-IF innovative technology allows you to stain organoids and spheroids in just 8 hours, while still in hydrogel or extracellular matrix, preserving delicate structures and cellular integrity. This all-in-one reagent removes the need for harvesting, clearing, transferring and centrifuging, preventing sample loss and ensuring consistent accurate results.
Are you having issues with unsuccessful labelling of precious samples?
CellO-IF has been proven to help with labelling samples using primary antibodies that were unsuccessful with the traditional methods. Available in two formats for labelling both 3D and 2D samples producing high quality images.
Features
- High quality images in hours, not days
- Reduced number of reagents and steps
- Label organoids, spheroids, and cells while still in hydrogel or extracellular matrix
- Reduces background noise for high-resolution images
- Boosted antigenicity for clearer visualization of target
Benefits
- Faster Results
- Streamlined Workflows
- Effortless labelling
- Enhanced Clarity
- Optimize detection
Easy, Simple and fast protocol
1. Fix in 4% paraformaldehyde (PFA) for 1 hr at 37°C
2. Rinse with PBS, X3, 10 min at 37°C
3. Incubate for 1.5-2 hrs at 37 °C with primary antibodies diluted in CellO-IF
4. Wash with CellO-IF X3, 10 min at 37°C
5. Incubate for 1.5-2 hrs with secondary antibodies diluted in CellO-IF
6. Wash with PBS, X3, 10 min at 37°C
7. Mount & image under microscope
Important application notes:
- Incubation Conditions: Keeping the samples at 37°C during the experiments is critical for maintaining optimal conditions and getting the best results. Samples can be removed from the incubator for solution changes or antibody additions at room temperature but it is essential to return them to the incubator at 37°C immediately afterward.
- Overnight Incubation: It is important to note that leaving samples in CellO-IF (with primary antibodies) overnight can potentially damage the samples. We strongly recommend following the protocol's guidelines for incubation times. You may consider extending the incubation times slightly for larger samples but it is recommended to keep them within the recommended 2-3 hour range.
Request your CellO-IF Sample
Want to try before you buy? Click here to request your CellO-IF Sample! Please note there are limited numbers of samples, so we may not be able to provide all requests if there is high demand.
| Name | Datasheet | Packsize | Order |
|---|---|---|---|
| CellO-IF2 All-in-one IF labeling reagent for cells (15 mL) |  | 15 ml | View |
| CellO-IF2 All-in-one IF labeling reagent for cells (45 mL) | | 45 mL | View |
| CellO-IF2 All-in-one IF labeling reagent for cells (90 mL) | | 90 mL | View |
| CellO-IF3 All-in-one IF labeling reagent for spheroids/organoids (15 mL) | | 15 ml | View |
| CellO-IF3 All-in-one IF labeling reagent for spheroids/organoids (45 mL) | | 45 mL | View |
| CellO-IF3 All-in-one IF labeling reagent for spheroids/organoids (90 mL) | | 90 mL | View |
Application Spotlight and Customer Reviews
CellO-IF can be used for a variety of 2D and 3D cell culture applications.
Neuronal Models
"We really didn’t think imaging the neuronal organoids would be as easy because they are so fragile and difficult to section with traditional methods. Using CellO™-IF simplified our workflow and it has great potential to make high-throughput imaging of organoids feasible. We got beautiful images using CellO-IF on both 2D and 3D cultures.”
“We could visualize POMC in primary neurons for the first time. The process is so simple and fast. ”
Staining of Liver Cancer Spheroids in just 6 hours!
Human lung organoid
“It is such a smart and simple technology for organoid studies. We could protect delicate patients’ samples during the entire process and get excellent results."
Frequently Asked Questions
Does the size of the hydrogel dome affect the success of labeling?
While smaller hydrogel domes often result in better reagent penetration and brighter images, larger domes can be successfully labelled with a slight adjustment. For larger volumes, chilling the sample on ice for 2 minutes before fixation can help reduce the hydrogel's thickness and improve reagent penetration.
What type of mounting medium do you recommend?
Glycerol or commercially available aqueous mounting media are suitable for mounting labeled samples. For nuclear labeling, consider a custom mounting medium containing Hoechst or DAPI.
Here’s a recipe of a mounting medium that also labels nuclei: 1,2 uL Hoechst or DAPI+675 uL PBS + 675 uL Glycerol
How much solution do I need for each sample?
The required solution volume varies based on the plate type and well size. Ensure the solution completely covers the sample. Here's a general guideline:
•24-well plate: 100-200 µL per well
•96-well plate: 50-100 µL per well
•Glass-bottomed dish: 150-200 µL
•CellO-M: 150-200 µL
Can we use this protocol for organoids that are not in a gel?
Yes, our protocol can be adapted for organoids that are not embedded in a gel.
Can I leave the sample in CellO-IF for an extended period, for example, overnight?
No, we DO NOT recommend leaving the samples in CellO-IF.
What should be the concentration of primary/secondary antibodies?
•The recommended dilutions from the manufacturer for both primary and secondary antibodies work for many experiments. The user may need to optimize some antibodies' dilution rates as in conventional methods. Some users prefer starting with two times higher dilution for primary antibodies to save time and their precious organoids.
CellO-IF also helps with labelling samples using primary antibodies that were unsuccessful with the traditional methods.
CellO-IF technology enhances labeling and eliminates the need for an antigen retrieval step.
What should be the incubation period with primary/secondary antibodies?
The incubation period with primary antibody may need to be optimized due to variations in the sample size, the antibody's feature, and the protein's location. Suppose the protein you want to visualize is on the cell membrane. In that case, the incubation period should be around 1hour for most of the primary antibodies. If the protein is in the cytoplasm, the incubation period changes between 1hr-1,5 hr. If the protein is in the nuclei, you may extend the incubation period to 2 hours. For multiple labeling of the proteins in the nuclei and the cell membrane, the user needs to optimize that duration between 1,5 to 2 hours.
How can I keep the sample temperature at 37°C during the entire protocol?
Here are several options:
•Place the sample in the incubator immediately after every step and be cautious not to leave the sample outside of the incubator for more than a few minutes.
•Leave your cell culture dish on a prewarmed hot plate at 37C during the experiment.
•Sterilize a steel block, warm it to 37C in the incubator before the experiment, replace it in the laminar flow, and then put the cell culture dish on this block during the experiment.
•Work in a laminar flow with a heated working space.
