Organoid Immune Co-culture

In recent years, cancer immunotherapy has become extremely popular in the treatment of cancer. The goal of tumor immunotherapy is to  stimulate or mobilize the body's immune system to enhance the anti-tumor immunity within the tumor microenvironment, thereby controlling and killing cancer cells. The rise of immunotherapy has also spurred the development of cancer research. However, due to the complexity of humanized systems and the partial or ineffective reconstitution of the immune system, immuno-oncology models face significant challenges. There is an urgent clinical need for in vitro models that can be used to verify the efficacy of personalized treatments. The advent of organoids has provided new opportunities for tumor immunotherapy, but the lack of immune cells in existing organoids has limited their development.

 

The method of tumor organoid immune co-culture is as follow：

1. Organoid Preparation

(1) One week in advance, revive and culture the organoids to a good condition;

(2) Take an appropriate number of organoids and wash them several times with pre-chilled PBS to remove most of the matrix gel;

(3) Resuspend the organoids in culture medium.

 

2. PBMC Separation and Organoid Co-culture

1. Peripheral blood is collected in a sterile environment using a vacuum blood collection tube containing EDTA-K2, and then transported and stored;

2. Fresh venous blood is diluted with PBS buffer solution at a 1:1 ratio and gently mixed for later use;

3. Add 4ml of cell separation liquid (abs930) to the 8ml venous blood mixture slowly and let it stand;

4. Centrifuge at 2000rpm for 20 minutes, during which the acceleration and deceleration speeds are reduced to 1 to prevent cell fusion;

5. At this point, observe that the 15ml centrifuge tube has four layers and gently collect the white suspension in the second layer into a 15ml centrifuge tube, mix well and set aside for use;

6. Centrifuge at 1500rpm for 10 minutes, during which the acceleration and deceleration speeds are reduced to 1 to prevent cell fusion, and discard the supernatant;

7. Add an appropriate amount of T cell serum-free base medium and let it stand for observation. Preparation of T cell serum-free complete medium: Add IL-2 at 200-500 units/ml (to stimulate T cell growth and proliferation) and CD3/CD28 magnetic beads at 3ug/ml (to activate T cells) to the T cell serum-free base medium;

8. Using the T cell serum-free complete medium, adjust the suspension to the appropriate concentration, and add approximately 5*10^5 viable T cells per well in a 96-well plate, with a volume of 100ul;

9. Resuspend the organoid mixture and add an equal amount of the sample to each well in the 96-well plate, to a total volume of 200ul.

 

3. Cultivation and Observation

(1) On D0, observe and record the status of organoids and T cells;

(2) Culture under conditions of 37℃ and 5% CO2, with daily continuous observation;

(3) On D3, remove 100ul of culture supernatant, add 100ul of fresh culture medium, and simultaneously add (treatment antibodies or drugs) to achieve a final concentration of 100nM (if significant differences are already observed on D3, record the morphology in a large field bright field and terminate the experiment);

(4) Continue observation until significant differences are observed between experimental groups, then take photos to record the morphology in a large field bright field;

(5) Preserve the cell supernatant for subsequent testing.

This figure represents the immune co-culture experiment conducted by our company. It is evident from the above image that under the influence of Drug B, T cells exhibit a significant cytotoxic effect.

 

 

Item NO.	Product Name	Size
abs9514	Mouse Intestinal Organoid Culture Medium Kit	1kit
abs9516	Mouse Liver Organoid Culture Medium Kit	1kit
abs9515	Mouse Lung Organoid Culture Medium Kit	1kit
abs9446	Human Breast Cancer Organoid Culture Medium Kit	1kit
abs9448	Human Endometrial carcinoma Organoid Culture Medium Kit	1kit
abs9445	Human Colorectal cancer Organoid Culture Medium Kit	1kit
abs9443	Human Lung Cancer Organoid Culture Medium Kit	1kit
abs9449	Human Gastric Cancer Organoid Culture Medium Kit	1kit
abs9447	Human Pancreatic Cancer Organoid Culture Medium Kit	1kit
abs9558	Human Ovarian Cancer Organoid Culture Medium Kit	1kit
abs9544	Human Liver Organoid Culture Medium Kit	1kit
abs9546	Human Lung Organoid Culture Medium Kit	1kit
abs9549	Mouse Gastric Cancer Organoid Culture Medium Kit	1kit
abs9550	Mouse Lung Cancer Organoid Culture Medium Kit	1kit
abs9551	Mouse Breast Cancer Organoid Culture Medium Kit	1kit
abs9552	Mouse Hepatocarcinoma Organoid Culture Medium Kit	1kit
abs9553	Mouse Pancreatic Cancer Organoid Culture Medium Kit	1kit

 

Absin provides antibodies, proteins, ELISA kits, cell culture, detection kits, and other research reagents. If you have any product needs, please contact us.

Absin Bioscience Inc. Email: worldwide@absin.cn

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