bioGenous肿瘤器官基础培养基 Cancer Organoid Basal Medium(B213152)

肿瘤器官基础培养基 Cancer Organoid Basal Medium

货号:B213152

规格:500ml

品牌:bioGenous

产品介绍

Product Description

bioGenousTM Cancer Organoid Basal Medium is a basal medium optimized for the culture of mammalian cancer organoids. Cancer Organoid Basal Medium requires supplementation with serum replacements, typically bioGenousTM B plus supplement (B846028) and growth factors, since this medium does not contain proteins and any other animal origin components.

 


技术参数

Product Information

Component

Component Cat#

Volume

Storage & Stability

bioGenousTM Cancer Organoid Basal Medium(50x)

B213152

500mL

2-8 °C, protected from strong light,

12 months

Specifications

Purification

Sterility Testing

Shipping Conditions

Applications

Animal Origin

Country of Origin

0.2 μm filtered

Pass

Ambient

Organoid Culture

None

China

Main Additives

With

Without

D-Glucose

L-glutamine

Non Essential Amino Acids

HEPES

Sodium Pyruvate

Ascorbic Acid

Niacinamide

Folic Acid

Choline chloride

i-Inositol

Glutathione

Phenol Red

NOTE: Cancer Organoid Basal Medium uses a sodium bicarbonate buffer system and therefore requires a 5–10% CO2 environment to maintain physiological pH. Always use aseptic techniques when handling and supplementing Cancer Organoid Basal Medium.

Storege and Handling

Cancer Organoid Basal Medium is supplied in gamma-irradiated, sterile PETG or PETE bottles.It is recommended to be stored at a temperature of 2-8 °C, and protected from strong light.

Precaution:

When handling bio-hazardous materials such as human cells, safe laboratory procedures should be followed, and personal protective equipment should be worn.

cGMP Manufacturing and Quality System

Cancer Organoid Basal Medium is manufactured in cGMP-compliant facilities certified with ISO9001ISO14000ISO13485 quality system standards.

Limitations

FOR LABORATORY RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES.

bioGenous宫颈癌 Cervical Cancer Organoid Kit(K2169-CC)

宫颈癌 Cervical Cancer Organoid Kit

货号:K2169-CC

规格:100ml

500ml

品牌:bioGenous

产品介绍

Product Description:

bioGenousTM Cervical Cancer Organoid Kit is a chemically defined cell culture medium for the establishment and maintenance of human cervical cancer organoids. Patient-derived cancer organoids recapitulate the genomic and pathological features of original tumors and therefore hold great promise for medical research and precision medicine.


技术参数

Product Information:

Component

Component Cat#

Volume

Storage & Stability

bioGenousTM Cervical Cancer Organoid Basal Medium

K2169-CC-A100/A500

100mL/500mL

4℃,12 months

bioGenousTM Cervical Cancer Organoid Supplement B (50x)

K2169-CC-B100/B500

2mL/10mL

-20, avoid repeated freeze-thaw cycles, 12 months

bioGenousTM Cervical Cancer Organoid Supplement C (250x)

K2169-CC-C100/C500

0.4mL/2mL

-20, avoid repeated freeze-thaw cycles, 12 months

Preparation of Cervical Cancer Organoid Complete Medium

Use a sterile technique to prepare the cervical cancer organoid complete medium. The following example is for preparing 10mL complete medium. If preparing other volumes, adjust accordingly.

1.      Thaw Cervical Cancer Organoid Supplement B(50x) and Cervical Cancer Organoid Supplement C(250x) on ice. Mix thoroughly.

NOTE: Once thawed, use immediately or aliquot and store at -20°C for not more than 10 months. After thawing the aliquots, use immediately. Do not re-freeze.

2.      Add 200μL Cervical Cancer Organoid Supplement B(50x) and 40μL Cervical Cancer Organoid Supplement C(250x) to 9.76mL Cervical Cancer Organoid Basal Medium. Mix thoroughly.

NOTE: If not used immediately, store the complete medium at 2-8°C for not more than 2 weeks. The Cervical Cancer Organoid Supplement B contains fungicide and antibiotics (50x).

Protocol for Establishment of Patient-Derived Cervical Cancer Organoids

CAUTION Studies involving primary human tissue material must follow all relevant institutional and government regulations. Informed consent must be obtained from all subjects before the collection of the primary human tissue material.

Establishment of Organoids from Primary Tissue

1.      Collect primary human cervical cancer tissue pieces in ice-cold Primary Tissue Storage Solution (K601005) with conical tubes. Keep tissue samples at 4°C until the start of the isolation.

2.      Assess whether the obtained tissue pieces consist purely of epithelium. If fat or muscle tissues are present, remove these non-epithelial components as much as possible using surgical scissors or scalpels and forceps under a dissection microscope. If no fat or muscle tissues are present, continue to the next step immediately.

3.      Rinse the tissue with Cancer Organoid Basal Medium (B213152) or DPBS twice.

4.      Mince the tissue into small fragments of 1-3 mm3 in a cell culture dish using surgical scissors or scalpels.

5.      Digest the tissue fragments with 10mL of Tumor Tissue Digestion Solution (K601003) in a 15mL conical tube at 37°C, with variable incubation times ranging from 30 min to 1 h. Carefully monitor the digestion process, mixing the content of the tube every 5-10 min by shaking vigorously or pipetting the mixture up and down. The digestion process could be finished when most of tissue fragments are able to pass through the 1mL pipette tips.

6.      Add FBS to the tissue digestion mixture to a final concentration of 2%, and filter using a 100 μm cell strainer.

7.      Collect and centrifuge the filtered cells at 250g for 3 min at 4 °C. In case of a visible red pellet, aspirate the supernatant, and resuspend the pellet using 2mL of Red Blood Cell Lysis Solution (E238010) to lyse the erythrocytes at room temperature for 1 min and centrifuge at 250g for 3 min at 4°C.

8.      Aspirate the supernatant and resuspend the pellet in Cancer Organoid Basal Medium, centrifuge at 250g for 3 min at 4°C, repeat this step once more time.

9.      Aspirate the supernatant and resuspend the pellet in Matrigel. The Matrigel should be kept on ice to prevent it from solidifying. Perform the process as quickly as possible. The amount of Matrigel used depends on the size of the pellet. Approximately 10,000 cells should be plated in 25 μL of Matrigel.

CRITICAL: Do not overly dilute the Matrigel (Matrigel should be >70% (Matrigel vol/Total vol)), as this may inhibit the proper formation of the solid droplets.

10.   Plate the Matrigel containing organoids at the bottom of 24-well cell culture plates in droplets of ~30 μL each around the center of the well..

CRITICAL: Once the organoids are resuspended in Matrigel, proceed with plating as quickly as possible, as the Matrigel may solidify in the tube or pipette tip. Do not let the Matrigel touch the wall of wells.

11.   Place the culture plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15-25 min to let the Matrigel solidify.

12.   Prepare the required amount of cervical cancer organoid complete medium.

13.   Once the Matrigel droplets have solidified (15-25 min), open the plate and carefully add 500 μL of organoid complete medium to each well.

CRITICAL: Do not add the medium directly on top of the Matrigel droplets, as this might disrupt the droplets.

14.   Place the culture plate in a humidified incubator at 37 °C and 5% (vol/vol) CO2.

15.   Change the medium every 3-4 d by carefully aspirating the medium from the wells and replacing it with a fresh, pre-warmed organoid complete medium.

16.   Closely monitor the organoid formation. Ideally, patient-derived cervical cancer organoids should be passaged for the first time between 7 and 10 d after the initial plating.

Splitting and Passaging of Organoids

17.   Pipette up and down to disrupt the Matrigel and transfer the organoid suspension to a 1.5 mL conical tube.

18.   Pipette the organoid suspension up and down to mix thoroughly by pipetting against the bottom of the tube to create pressure, which will aid the removal of Matrigel.

19.   Centrifuge organoids at 250g for 3 min at room temperature.

20.   Aspirate the supernatant and split organoids using either Organoid Dissociation Solution (E238001) or by mechanical disruption. For Organoid Dissociation Solution-based cell dissociation, resuspend the pellet in 0.2 mL of Organoid Dissociation Solution, pipette up and down and incubate at 37 °C until organoids fall apart. Pipette up and down with a filter tip for ≥8 times every 2 min to aid in the disruption of the organoids. Closely monitor the digestion to keep the incubation time in the Organoid Dissociation Solution to a minimum. In case of mechanical disruption, resuspend the pellet in 1.5 mL of Cancer Organoid Basal Medium. Carefully pipette the organoid suspension up and down 30 times by pipetting against the bottom of the tube to create pressure, which will aid organoid disruption.

CRITICAL: Do not dissociate in Organoid Dissociation Solution for >7 min, as this may result in poor organoid outgrowth or even loss of the culture. As a rule of thumb, digestion is complete if a mixture of small lumps of cells (consisting of 10–50 cells) can be observed.

21.   After shearing is complete, wash once by topping up with 1 mL of Cancer Organoid Basal Medium, and centrifuge at 250g for 3 min at room temperature.

22.   Aspirate the supernatant and resuspend the organoid pellet in 70% (vol/vol) Matrigel, and plate organoids in droplets at the bottom of a culture plate as described in Steps 10. After plating is complete, transfer the plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15–25 min.

23.   Pre-warm cervical cancer organoid complete medium at 37 °C.

24.   After the Matrigel droplets have solidified (15–25 min), carefully pipette pre-warmed medium into the wells.

25.   Place culture plates in a humidified incubator at 37 °C and 5% (vol/vol) CO2 until the organoids are needed for further experiments.

bioGenous结直肠癌 Colorectal Cancer Organoid Kit(K2103-CR)

结直肠癌 Colorectal Cancer Organoid Kit

货号:K2103-CR

规格:100ml

500ml

品牌:bioGenous

产品介绍

Product Description:

bioGenousTM Colorectal Cancer Organoid Kit is a chemically defined cell culture medium for the establishment and maintenance of human Colorectal cancer organoids. Patient-derived cancer organoids recapitulate the genomic and pathological features of original tumors and therefore hold great promise for medical research and precision medicine.


技术参数

Product Information:

Component

Component Cat#

Volume

Storage& Stability

bioGenousTM Colorectal Cancer Organoid Basal Medium

K2103-CR-A100/A500

100mL/500mL

4℃,12 months

bioGenousTM Colorectal Cancer Organoid Supplement B (50x)

K2103-CR-B100/B500

2mL/10mL

-20,avoid repeated freeze-thaw cycles, 12 months

bioGenousTM Colorectal Cancer Organoid Supplement C (250x)

K2103-CR-C100/C500

0.4mL/2mL

-20, avoid repeated freeze-thaw cycles, 12 months

Preparation of Colorectal Cancer Organoid Complete Medium

Use a sterile technique to prepare the colorectal cancer organoid complete medium. The following example is for preparing 10mL complete medium. If preparing other volumes, adjust accordingly.

1.       Thaw Colorectal Cancer Organoid Supplement B(50x) and Colorectal Cancer Organoid Supplement C(250x) on ice. Mix thoroughly.

NOTE: Once thawed, use immediately or aliquot and store at -20°C for not more than 10 months. After thawing the aliquots, use immediately. Do not re-freeze.

2.       Add 200μL Colorectal Cancer Organoid Supplement B(50x) and 40μL Colorectal Cancer Organoid Supplement C(250x) to 9.76mL Colorectal Cancer Organoid Basal Medium. Mix thoroughly.

NOTE: If not used immediately, store the complete medium at 2-8°C for not more than 2 weeks. The Colorectal Cancer Organoid Supplement B contains fungicide and antibiotics (50x).

Protocol for Establishment of Patient-Derived Colorectal Cancer Organoids

CAUTIONStudies involving primary human tissue material must follow all relevant institutional and government regulations. Informed consent must be obtained from all subjects before the collection of the primary human tissue material.

Establishment of Organoids from Primary Tissue

1.       Collect primary human colorectal cancer tissue pieces in ice-cold Primary Tissue Storage Solution (K601005) with conical tubes. Keep tissue samples at 4°C until the start of the isolation.

2.       Assess whether the obtained tissue pieces consist purely of epithelium. If fat or muscle tissues are present, remove these non-epithelial components as much as possible using surgical scissors or scalpels and forceps under a dissection microscope. If no fat or muscle tissues are present, continue to the next step immediately.

3.       Rinse the tissue with Cancer Organoid Basal Medium (B213152) or DPBS twice.

4.       Mince the tissue into small fragments of 1-3 mm3 in a cell culture dish using surgical scissors or scalpels.

5.       Digest the tissue fragments with 10mL of Tumor Tissue Digestion Solution (K601003) in a 15mL conical tube at 37°C, with variable incubation times ranging from 30 min to 1.5 h. Carefully monitor the digestion process, mixing the content of the tube every 5-10 min by shaking vigorously or pipetting the mixture up and down. The digestion process could be finished when most of tissue fragments are able to pass through the 1mL pipette tips.

6.       Add FBS to the tissue digestion mixture to a final concentration of 2%, and filter using a 100 μm cell strainer.

7.       Collect and centrifuge the filtered cells at 250g for 3 min at 4 °C. In case of a visible red pellet, aspirate the supernatant, and resuspend the pellet using 2mL of Red Blood Cell Lysis Solution (E238010) to lyse the erythrocytes at room temperature for 1 min and centrifuge at 250g for 3 min at 4°C.

8.       Aspirate the supernatant and resuspend the pellet in Cancer Organoid Basal Medium, centrifuge at 250g for 3 min at 4°C, repeat this step once more time.

9.       Aspirate the supernatant and resuspend the pellet in Matrigel. The Matrigel should be kept on ice to prevent it from solidifying. Perform the process as quickly as possible. The amount of Matrigel used depends on the size of the pellet. Approximately 10,000 cells should be plated in 25 μL of Matrigel.

CRITICAL: Do not overly dilute the Matrigel (Matrigel should be >70% (Matrigel vol/Total vol)), as this may inhibit the proper formation of the solid droplets.

10.     Plate the Matrigel containing organoids at the bottom of 24-well cell culture plates in droplets of ~30 μL each around the center of the well..

CRITICAL: Once the organoids are resuspended in Matrigel, proceed with plating as quickly as possible, as the Matrigel may solidify in the tube or pipette tip. Do not let the Matrigel touch the wall of wells.

11.     Place the culture plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15-25 min to let the Matrigel solidify.

12.     Prepare the required amount of colorectal cancer organoid complete medium.

13.     Once the Matrigel droplets have solidified (15-25 min), open the plate and carefully add 500 μL of organoid complete medium to each well.

CRITICAL: Do not add the medium directly on top of the Matrigel droplets, as this might disrupt the droplets.

14.     Place the culture plate in a humidified incubator at 37 °C and 5% (vol/vol) CO2.

15.     Change the medium every 3-4 d by carefully aspirating the medium from the wells and replacing it with a fresh, pre-warmed organoid complete medium.

16.     Closely monitor the organoid formation. Ideally, patient-derived colorectal cancer organoids should be passaged for the first time between 7 and 10 d after the initial plating.

Splitting and Passaging of Organoids

17.     Pipette up and down to disrupt the Matrigel and transfer the organoid suspension to a 1.5 mL conical tube.

18.     Pipette the organoid suspension up and down to mix thoroughly by pipetting against the bottom of the tube to create pressure, which will aid the removal of Matrigel.

19.     Centrifuge organoids at 250g for 3 min at room temperature.

20.     Aspirate the supernatant and split organoids using either Organoid Dissociation Solution (E238001) or by mechanical disruption. For Organoid Dissociation Solution-based cell dissociation, resuspend the pellet in 0.2 mL of Organoid Dissociation Solution, pipette up and down and incubate at 37 °C until organoids fall apart. Pipette up and down with a filter tip for ≥8 times every 2 min to aid in the disruption of the organoids. Closely monitor the digestion to keep the incubation time in the Organoid Dissociation Solution to a minimum. In case of mechanical disruption, resuspend the pellet in 1.5 mL of Cancer Organoid Basal Medium. Carefully pipette the organoid suspension up and down 30 times by pipetting against the bottom of the tube to create pressure, which will aid organoid disruption.

CRITICAL: Do not dissociate in Organoid Dissociation Solution for >6 min, as this may result in poor organoid outgrowth or even loss of the culture. As a rule of thumb, digestion is complete if a mixture of small lumps of cells (consisting of 10–50 cells) can be observed.

21.     After shearing is complete, wash once by topping up with 1 mL ofCancer Organoid Basal Medium, and centrifuge at 250g for 3 min at room temperature.

22.     Aspirate the supernatant and resuspend the organoid pellet in 70% (vol/vol) Matrigel, and plate organoids in droplets at the bottom of a culture plate as described in Steps 10. After plating is complete, transfer the plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15–25 min.

23.     Pre-warm colorectal cancer organoid complete medium at 37 °C.

24.     After the Matrigel droplets have solidified (15–25 min), carefully pipette pre-warmed medium into the wells.

Place culture plates in a humidified incubator at 37 °C and 5% (vol/vol) CO2 until the organoids are needed for further experiments.

bioGenous卵巢癌 Ovarian Cancer Organoid Kit(K2168-OC)

卵巢癌 Ovarian Cancer Organoid Kit

货号:K2168-OC

规格:100ml

500ml

品牌:bioGenous

产品介绍

Product Description:

bioGenousTM Ovarian Cancer Organoid Kit is a chemically defined cell culture medium for the establishment and maintenance of human Ovarian Cancer organoids. Patient-derived cancer organoids recapitulate the genomic and pathological features of original tumors and therefore hold great promise for medical research and precision medicine.


技术参数

Product Information:

Component

Component Cat#

Volume

Storage & Stability

bioGenousTM Ovarian Cancer Organoid Basal Medium

K2168-OC-A100/A500

100mL/500mL

4℃,12 months

bioGenousTM Ovarian Cancer Organoid Supplement B (50x)

K2168-OC-B100/B500

2mL/10mL

-20, avoid repeated freeze-thaw cycles, 12 months

bioGenousTM Ovarian Cancer Organoid Supplement C (250x)

K2168-OC-C100/C500

0.4mL/2mL

-20, avoid repeated freeze-thaw cycles, 12 months

Preparation of Ovarian Cancer Organoid Complete Medium

Use a sterile technique to prepare the ovarian cancer organoid complete medium. The following example is for preparing 10mL complete medium. If preparing other volumes, adjust accordingly.

1.      Thaw Ovarian Cancer Organoid Supplement B(50x) and Ovarian Cancer Organoid Supplement C(250x) on ice. Mix thoroughly.

NOTE: Once thawed, use immediately or aliquot and store at -20°C for not more than 10 months. After thawing the aliquots, use immediately. Do not re-freeze.

2.      Add 200μL Ovarian Cancer Organoid Supplement B(50x) and 40μL Ovarian Cancer Organoid Supplement C(250x) to 9.76mL Ovarian Cancer Organoid Basal Medium. Mix thoroughly.

NOTE: If not used immediately, store the complete medium at 2-8°C for not more than 2 weeks. The Ovarian Cancer Organoid Supplement B contains fungicide and antibiotics (50x).

Protocol for Establishment of Patient-Derived Ovarian Cancer Organoids

CAUTION Studies involving primary human tissue material must follow all relevant institutional and government regulations. Informed consent must be obtained from all subjects before the collection of the primary human tissue material.

Establishment of Organoids from Primary Tissue

1.      Collect primary human ovarian cancer tissue pieces in ice-cold Primary Tissue Storage Solution (K601005) with conical tubes. Keep tissue samples at 4°C until the start of the isolation.

2.      Assess whether the obtained tissue pieces consist purely of epithelium. If fat or muscle tissues are present, remove these non-epithelial components as much as possible using surgical scissors or scalpels and forceps under a dissection microscope. If no fat or muscle tissues are present, continue to the next step immediately.

3.      Rinse the tissue with Cancer Organoid Basal Medium (B213152) or DPBS twice.

4.      Mince the tissue into small fragments of 1-3 mm3 in a cell culture dish using surgical scissors or scalpels.

5.      Digest the tissue fragments with 10mL of Tumor Tissue Digestion Solution (K601003) in a 15mL conical tube at 37°C, with variable incubation times ranging from 30 min to 1.5 h. Carefully monitor the digestion process, mixing the content of the tube every 5-10 min by shaking vigorously or pipetting the mixture up and down. The digestion process could be finished when most of tissue fragments are able to pass through the 1mL pipette tips.

6.      Add FBS to the tissue digestion mixture to a final concentration of 2%, and filter using a 100 μm cell strainer.

7.      Collect and centrifuge the filtered cells at 250g for 3 min at 4 °C. In case of a visible red pellet, aspirate the supernatant, and resuspend the pellet using 2mL of Red Blood Cell Lysis Solution (E238010) to lyse the erythrocytes at room temperature for 1 min and centrifuge at 250g for 3 min at 4°C.

8.      Aspirate the supernatant and resuspend the pellet in Cancer Organoid Basal Medium, centrifuge at 250g for 3 min at 4°C, repeat this step once more time.

9.      Aspirate the supernatant and resuspend the pellet in Matrigel. The Matrigel should be kept on ice to prevent it from solidifying. Perform the process as quickly as possible. The amount of Matrigel used depends on the size of the pellet. Approximately 10,000 cells should be plated in 25 μL of Matrigel.

CRITICAL: Do not overly dilute the Matrigel (Matrigel should be >70% (Matrigel vol/Total vol)), as this may inhibit the proper formation of the solid droplets.

10.   Plate the Matrigel containing organoids at the bottom of 24-well cell culture plates in droplets of ~30 μL each around the center of the well..

CRITICAL: Once the organoids are resuspended in Matrigel, proceed with plating as quickly as possible, as the Matrigel may solidify in the tube or pipette tip. Do not let the Matrigel touch the wall of wells.

11.   Place the culture plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15-25 min to let the Matrigel solidify.

12.   Prepare the required amount of ovarian cancer organoid complete medium.

13.   Once the Matrigel droplets have solidified (15-25 min), open the plate and carefully add 500 μL of organoid complete medium to each well.

CRITICAL: Do not add the medium directly on top of the Matrigel droplets, as this might disrupt the droplets.

14.   Place the culture plate in a humidified incubator at 37 °C and 5% (vol/vol) CO2.

15.   Change the medium every 3-4 d by carefully aspirating the medium from the wells and replacing it with a fresh, pre-warmed organoid complete medium.

16.   Closely monitor the organoid formation. Ideally, patient-derived ovarian cancer organoids should be passaged for the first time between 7 and 10 d after the initial plating.

Splitting and Passaging of Organoids

17.   Pipette up and down to disrupt the Matrigel and transfer the organoid suspension to a 1.5 mL conical tube.

18.   Pipette the organoid suspension up and down to mix thoroughly by pipetting against the bottom of the tube to create pressure, which will aid the removal of Matrigel.

19.   Centrifuge organoids at 250g for 3 min at room temperature.

20.   Aspirate the supernatant and split organoids using either Organoid Dissociation Solution (E238001) or by mechanical disruption. For Organoid Dissociation Solution-based cell dissociation, resuspend the pellet in 0.2 mL of Organoid Dissociation Solution, pipette up and down and incubate at 37 °C until organoids fall apart. Pipette up and down with a filter tip for ≥8 times every 2 min to aid in the disruption of the organoids. Closely monitor the digestion to keep the incubation time in the Organoid Dissociation Solution to a minimum. In case of mechanical disruption, resuspend the pellet in 1.5 mL of Cancer Organoid Basal Medium. Carefully pipette the organoid suspension up and down 30 times by pipetting against the bottom of the tube to create pressure, which will aid organoid disruption.

CRITICAL: Do not dissociate in Organoid Dissociation Solution for >7 min, as this may result in poor organoid outgrowth or even loss of the culture. As a rule of thumb, digestion is complete if a mixture of small lumps of cells (consisting of 10–50 cells) can be observed.

21.   After shearing is complete, wash once by topping up with 1 mL of Cancer Organoid Basal Medium, and centrifuge at 250g for 3 min at room temperature.

22.   Aspirate the supernatant and resuspend the organoid pellet in 70% (vol/vol) Matrigel, and plate organoids in droplets at the bottom of a culture plate as described in Steps 10. After plating is complete, transfer the plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15–25 min.

23.   Pre-warm ovarian cancer organoid complete medium at 37 °C.

24.   After the Matrigel droplets have solidified (15–25 min), carefully pipette pre-warmed medium into the wells.

25.   Place culture plates in a humidified incubator at 37 °C and 5% (vol/vol) CO2 until the organoids are needed for further experiments.

bioGenous乳腺癌 Breast Cancer Organoid Kit(K2147-BC)

乳腺癌 Breast Cancer Organoid Kit

货号:K2147-BC

规格:100ml

500ml

品牌:bioGenous

产品介绍

Product Description:

bioGenousTM Breast Cancer Organoid Kit is a chemically defined cell culture medium for the establishment and maintenance of human breast cancer organoids. Patient-derived cancer organoids recapitulate the genomic and pathological features of original tumors and therefore hold great promise for medical research and precision medicine.


技术参数

Product Information:

Component

Component Cat#

Volume

Storage & Stability

bioGenousTM Breast Cancer Organoid Basal Medium

K2147-BC-A100/A500

100mL/500mL

4℃,12 months

bioGenousTM Breast Cancer Organoid Supplement B (50x)

K2147-BC-B100/B500

2mL/10mL

-20, avoid repeated freeze-thaw cycles, 12 months

bioGenousTM Breast Cancer Organoid Supplement C (250x)

K2147-BC-C100/C500

0.4mL/2mL

-20, avoid repeated freeze-thaw cycles, 12 months

Preparation of Breast Cancer Organoid Complete Medium

Use a sterile technique to prepare the breast cancer organoid complete medium. The following example is for preparing 10mL complete medium. If preparing other volumes, adjust accordingly.

1.      Thaw Breast Cancer Organoid Supplement B(50x) and Breast Cancer Organoid Supplement C(250x) on ice. Mix thoroughly.

NOTE: Once thawed, use immediately or aliquot and store at -20°C for not more than 10 months. After thawing the aliquots, use immediately. Do not re-freeze.

2.      Add 200μL Breast Cancer Organoid Supplement B(50x) and 40μL Breast Cancer Organoid Supplement C(250x) to 9.76mL Breast Cancer Organoid Basal Medium. Mix thoroughly.

NOTE: If not used immediately, store the complete medium at 2-8°C for not more than 2 weeks. The Breast Cancer Organoid Supplement B contains fungicide and antibiotics (50x).

Protocol for Establishment of Patient-Derived Breast Cancer Organoids

CAUTION Studies involving primary human tissue material must follow all relevant institutional and government regulations. Informed consent must be obtained from all subjects before the collection of the primary human tissue material.

Establishment of Organoids from Primary Tissue

1.      Collect primary human breast cancer tissue pieces in ice-cold Primary Tissue Storage Solution (K601005) with conical tubes. Keep tissue samples at 4°C until the start of the isolation.

2.      Assess whether the obtained tissue pieces consist purely of epithelium. If fat or muscle tissues are present, remove these non-epithelial components as much as possible using surgical scissors or scalpels and forceps under a dissection microscope. If no fat or muscle tissues are present, continue to the next step immediately.

3.      Rinse the tissue with Cancer Organoid Basal Medium (B213152) or DPBS twice.

4.      Mince the tissue into small fragments of 1-3 mm3 in a cell culture dish using surgical scissors or scalpels.

5.      Digest the tissue fragments with 10mL of Tumor Tissue Digestion Solution (K601003) in a 15mL conical tube at 37°C, with variable incubation times ranging from 30 min to 1.5 h. Carefully monitor the digestion process, mixing the content of the tube every 5-10 min by shaking vigorously or pipetting the mixture up and down. The digestion process could be finished when most of tissue fragments are able to pass through the 1mL pipette tips.

6.      Add FBS to the tissue digestion mixture to a final concentration of 2%, and filter using a 100 μm cell strainer.

7.      Collect and centrifuge the filtered cells at 250g for 3 min at 4 °C. In case of a visible red pellet, aspirate the supernatant, and resuspend the pellet using 2mL of Red Blood Cell Lysis Solution (E238010) to lyse the erythrocytes at room temperature for 1 min and centrifuge at 250g for 3 min at 4°C.

8.      Aspirate the supernatant and resuspend the pellet in Cancer Organoid Basal Medium, centrifuge at 250g for 3 min at 4°C, repeat this step once more time.

9.      Aspirate the supernatant and resuspend the pellet in Matrigel. The Matrigel should be kept on ice to prevent it from solidifying. Perform the process as quickly as possible. The amount of Matrigel used depends on the size of the pellet. Approximately 10,000 cells should be plated in 25 μL of Matrigel.

CRITICAL: Do not overly dilute the Matrigel (Matrigel should be >70% (Matrigel vol/Total vol)), as this may inhibit the proper formation of the solid droplets.

10.   Plate the Matrigel containing organoids at the bottom of 24-well cell culture plates in droplets of ~30 μL each around the center of the well..

CRITICAL: Once the organoids are resuspended in Matrigel, proceed with plating as quickly as possible, as the Matrigel may solidify in the tube or pipette tip. Do not let the Matrigel touch the wall of wells.

11.   Place the culture plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15-25 min to let the Matrigel solidify.

12.   Prepare the required amount of breast cancer organoid complete medium.

13.   Once the Matrigel droplets have solidified (15-25 min), open the plate and carefully add 500 μL of organoid complete medium to each well.

CRITICAL: Do not add the medium directly on top of the Matrigel droplets, as this might disrupt the droplets.

14.   Place the culture plate in a humidified incubator at 37 °C and 5% (vol/vol) CO2.

15.   Change the medium every 3-4 d by carefully aspirating the medium from the wells and replacing it with a fresh, pre-warmed organoid complete medium.

16.   Closely monitor the organoid formation. Ideally, patient-derived breast cancer organoids should be passaged for the first time between 7 and 10 d after the initial plating.

Splitting and Passaging of Organoids

17.   Pipette up and down to disrupt the Matrigel and transfer the organoid suspension to a 1.5 mL conical tube.

18.   Pipette the organoid suspension up and down to mix thoroughly by pipetting against the bottom of the tube to create pressure, which will aid the removal of Matrigel.

19.   Centrifuge organoids at 250g for 3 min at room temperature.

20.   Aspirate the supernatant and split organoids using either Organoid Dissociation Solution (E238001) or by mechanical disruption. For Organoid Dissociation Solution-based cell dissociation, resuspend the pellet in 0.2 mL of Organoid Dissociation Solution, pipette up and down and incubate at 37 °C until organoids fall apart. Pipette up and down with a filter tip for ≥8 times every 2 min to aid in the disruption of the organoids. Closely monitor the digestion to keep the incubation time in the Organoid Dissociation Solution to a minimum. In case of mechanical disruption, resuspend the pellet in 1.5 mL of Cancer Organoid Basal Medium. Carefully pipette the organoid suspension up and down 30 times by pipetting against the bottom of the tube to create pressure, which will aid organoid disruption.

CRITICAL: Do not dissociate in Organoid Dissociation Solution for >7 min, as this may result in poor organoid outgrowth or even loss of the culture. As a rule of thumb, digestion is complete if a mixture of small lumps of cells (consisting of 10–50 cells) can be observed.

21.   After shearing is complete, wash once by topping up with 1 mL of Cancer Organoid Basal Medium, and centrifuge at 250g for 3 min at room temperature.

22.   Aspirate the supernatant and resuspend the organoid pellet in 70% (vol/vol) Matrigel, and plate organoids in droplets at the bottom of a culture plate as described in Steps 10. After plating is complete, transfer the plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15–25 min.

23.   Pre-warm breast cancer organoid complete medium at 37 °C.

24.   After the Matrigel droplets have solidified (15–25 min), carefully pipette pre-warmed medium into the wells.

25.   Place culture plates in a humidified incubator at 37 °C and 5% (vol/vol) CO2 until the organoids are needed for further experiments.

bioGenous食道癌 Esophageal Cancer Organoid Kit(K2177-ES)

食道癌 Esophageal Cancer Organoid Kit

货号:K2177-ES

规格:100ml

500ml

品牌:bioGenous

产品介绍

Product Description:

bioGenousTM Esophageal Cancer Organoid Kit is a chemically defined cell culture medium for the establishment and maintenance of human esophageal cancer organoids. Patient-derived cancer organoids recapitulate the genomic and pathological features of original tumors and therefore hold great promise for medical research and precision medicine.


技术参数

Product Information:

Component

Component Cat#

Volume

Storage & Stability

bioGenousTM Esophageal Cancer Organoid Basal Medium

K2177-ES-A100/A500

100mL/500mL

4℃,12 months

bioGenousTM Esophageal Cancer Organoid Supplement B (50x)

K2177-ES-B100/B500

2mL/10mL

-20, avoid repeated freeze-thaw cycles, 12 months

bioGenousTM Esophageal Cancer Organoid Supplement C (250x)

K2177-ES-C100/C500

0.4mL/2mL

-20, avoid repeated freeze-thaw cycles, 12 months

Preparation of Esophageal Cancer Organoid Complete Medium

Use a sterile technique to prepare the esophageal cancer organoid complete medium. The following example is for preparing 10mL complete medium. If preparing other volumes, adjust accordingly.

1.      Thaw Esophageal Cancer Organoid Supplement B(50x) and Esophageal Cancer Organoid Supplement C(250x) on ice. Mix thoroughly.

NOTE: Once thawed, use immediately or aliquot and store at -20°C for not more than 10 months. After thawing the aliquots, use immediately. Do not re-freeze.

2.      Add 200μL Esophageal Cancer Organoid Supplement B(50x) and 40μL Esophageal Cancer Organoid Supplement C(250x) to 9.76mL Esophageal Cancer Organoid Basal Medium. Mix thoroughly.

NOTE: If not used immediately, store the complete medium at 2-8°C for not more than 2 weeks. The Esophageal Cancer Organoid Supplement B contains fungicide and antibiotics (50x).

Protocol for Establishment of Patient-Derived Esophageal Cancer Organoids

CAUTION Studies involving primary human tissue material must follow all relevant institutional and government regulations. Informed consent must be obtained from all subjects before the collection of the primary human tissue material.

Establishment of Organoids from Primary Tissue

1.      Collect primary human esophageal cancer tissue pieces in ice-cold Primary Tissue Storage Solution (K601005) with conical tubes. Keep tissue samples at 4°C until the start of the isolation.

2.      Assess whether the obtained tissue pieces consist purely of epithelium. If fat or muscle tissues are present, remove these non-epithelial components as much as possible using surgical scissors or scalpels and forceps under a dissection microscope. If no fat or muscle tissues are present, continue to the next step immediately.

3.      Rinse the tissue with Cancer Organoid Basal Medium (B213152) or DPBS twice.

4.      Mince the tissue into small fragments of 1-3 mm3 in a cell culture dish using surgical scissors or scalpels.

5.      Digest the tissue fragments with 10mL of Tumor Tissue Digestion Solution (K601003) in a 15mL conical tube at 37°C, with variable incubation times ranging from 30 min to 1 h. Carefully monitor the digestion process, mixing the content of the tube every 5-10 min by shaking vigorously or pipetting the mixture up and down. The digestion process could be finished when most of tissue fragments are able to pass through the 1mL pipette tips.

6.      Add FBS to the tissue digestion mixture to a final concentration of 2%, and filter using a 100 μm cell strainer.

7.      Collect and centrifuge the filtered cells at 250g for 3 min at 4 °C. In case of a visible red pellet, aspirate the supernatant, and resuspend the pellet using 2mL of Red Blood Cell Lysis Solution (E238010) to lyse the erythrocytes at room temperature for 1 min and centrifuge at 250g for 3 min at 4°C.

8.      Aspirate the supernatant and resuspend the pellet in Cancer Organoid Basal Medium, centrifuge at 250g for 3 min at 4°C, repeat this step once more time.

9.      Aspirate the supernatant and resuspend the pellet in Matrigel. The Matrigel should be kept on ice to prevent it from solidifying. Perform the process as quickly as possible. The amount of Matrigel used depends on the size of the pellet. Approximately 10,000 cells should be plated in 25 μL of Matrigel.

CRITICAL: Do not overly dilute the Matrigel (Matrigel should be >70% (Matrigel vol/Total vol)), as this may inhibit the proper formation of the solid droplets.

10.   Plate the Matrigel containing organoids at the bottom of 24-well cell culture plates in droplets of ~30 μL each around the center of the well..

CRITICAL: Once the organoids are resuspended in Matrigel, proceed with plating as quickly as possible, as the Matrigel may solidify in the tube or pipette tip. Do not let the Matrigel touch the wall of wells.

11.   Place the culture plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15-25 min to let the Matrigel solidify.

12.   Prepare the required amount of esophageal cancer organoid complete medium.

13.   Once the Matrigel droplets have solidified (15-25 min), open the plate and carefully add 500 μL of organoid complete medium to each well.

CRITICAL: Do not add the medium directly on top of the Matrigel droplets, as this might disrupt the droplets.

14.   Place the culture plate in a humidified incubator at 37 °C and 5% (vol/vol) CO2.

15.   Change the medium every 3-4 d by carefully aspirating the medium from the wells and replacing it with a fresh, pre-warmed organoid complete medium.

16.   Closely monitor the organoid formation. Ideally, patient-derived esophageal cancer organoids should be passaged for the first time between 7 and 10 d after the initial plating.

Splitting and Passaging of Organoids

17.   Pipette up and down to disrupt the Matrigel and transfer the organoid suspension to a 1.5 mL conical tube.

18.   Pipette the organoid suspension up and down to mix thoroughly by pipetting against the bottom of the tube to create pressure, which will aid the removal of Matrigel.

19.   Centrifuge organoids at 250g for 3 min at room temperature.

20.   Aspirate the supernatant and split organoids using either Organoid Dissociation Solution (E238001) or by mechanical disruption. For Organoid Dissociation Solution-based cell dissociation, resuspend the pellet in 0.2 mL of Organoid Dissociation Solution, pipette up and down and incubate at 37 °C until organoids fall apart. Pipette up and down with a filter tip for ≥8 times every 2 min to aid in the disruption of the organoids. Closely monitor the digestion to keep the incubation time in the Organoid Dissociation Solution to a minimum. In case of mechanical disruption, resuspend the pellet in 1.5 mL of Cancer Organoid Basal Medium. Carefully pipette the organoid suspension up and down 30 times by pipetting against the bottom of the tube to create pressure, which will aid organoid disruption.

CRITICAL: Do not dissociate in Organoid Dissociation Solution for >7 min, as this may result in poor organoid outgrowth or even loss of the culture. As a rule of thumb, digestion is complete if a mixture of small lumps of cells (consisting of 10–50 cells) can be observed.

21.   After shearing is complete, wash once by topping up with 1 mL of Cancer Organoid Basal Medium, and centrifuge at 250g for 3 min at room temperature.

22.   Aspirate the supernatant and resuspend the organoid pellet in 70% (vol/vol) Matrigel, and plate organoids in droplets at the bottom of a culture plate as described in Steps 10. After plating is complete, transfer the plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15–25 min.

23.   Pre-warm esophageal cancer organoid complete medium at 37 °C.

24.   After the Matrigel droplets have solidified (15–25 min), carefully pipette pre-warmed medium into the wells.

Place culture plates in a humidified incubator at 37 °C and 5% (vol/vol) CO2 until the organoids are needed for further experiments.

bioGenous胃癌 Gastric Cancer Organoid Kit(K2179-GC)

胃癌 Gastric Cancer Organoid Kit

货号:K2179-GC

规格:100ml

500ml

品牌:bioGenous

产品介绍

Product Description:

bioGenousTM Gastric Cancer Organoid Kit is a chemically defined cell culture medium for the establishment and maintenance of human gastric cancer organoid s. Patient-derived cancer organoids recapitulate the genomic and pathological features of original tumors and therefore hold great promise for medical research and precision medicine.


技术参数

Product Information:

Component

Component Cat#

Volume

Storage & Stability

bioGenousTM Gastric Cancer Organoid Basal Medium

K2179-GC-A100/A500

100mL/500mL

4℃,12 months

bioGenousTM Gastric Cancer Organoid Supplement B (50x)

K2179-GC-B100/B500

2mL/10mL

-20, avoid repeated freeze-thaw cycles, 12 months

bioGenousTM Gastric Cancer Organoid Supplement C (250x)

K2179-GC-C100/C500

0.4mL/2mL

-20, avoid repeated freeze-thaw cycles, 12 months

Preparation of Gastric Cancer Organoid Complete Medium

Use a sterile technique to prepare the gastric cancer organoid complete medium. The following example is for preparing 10mL complete medium. If preparing other volumes, adjust accordingly.

1.      Thaw Gastric Cancer Organoid Supplement B(50x) and Gastric Cancer Organoid Supplement C(250x) on ice. Mix thoroughly.

NOTE: Once thawed, use immediately or aliquot and store at -20°C for not more than 10 months. After thawing the aliquots, use immediately. Do not re-freeze.

2.      Add 200μL Gastric Cancer Organoid Supplement B(50x) and 40μL Gastric Cancer Organoid Supplement C(250x) to 9.76mL Gastric Cancer Organoid Basal Medium. Mix thoroughly.

NOTE: If not used immediately, store the complete medium at 2-8°C for not more than 2 weeks. The Gastric Cancer Organoid Supplement B contains fungicide and antibiotics (50x).

Protocol for Establishment of Patient-Derived Gastric Cancer Organoids

CAUTION Studies involving primary human tissue material must follow all relevant institutional and government regulations. Informed consent must be obtained from all subjects before the collection of the primary human tissue material.

Establishment of Organoids from Primary Tissue

1.      Collect primary human gastric cancer tissue pieces in ice-cold Primary Tissue Storage Solution (K601005) with conical tubes. Keep tissue samples at 4°C until the start of the isolation.

2.      Assess whether the obtained tissue pieces consist purely of epithelium. If fat or muscle tissues are present, remove these non-epithelial components as much as possible using surgical scissors or scalpels and forceps under a dissection microscope. If no fat or muscle tissues are present, continue to the next step immediately.

3.      Rinse the tissue with Cancer Organoid Basal Medium (B213152) or DPBS twice.

4.      Mince the tissue into small fragments of 1-3 mm3 in a cell culture dish using surgical scissors or scalpels.

5.      Digest the tissue fragments with 10mL of Tumor Tissue Digestion Solution (K601003) in a 15mL conical tube at 37°C, with variable incubation times ranging from 30 min to 1 h. Carefully monitor the digestion process, mixing the content of the tube every 5-10 min by shaking vigorously or pipetting the mixture up and down. The digestion process could be finished when most of tissue fragments are able to pass through the 1mL pipette tips.

6.      Add FBS to the tissue digestion mixture to a final concentration of 2%, and filter using a 100 μm cell strainer.

7.      Collect and centrifuge the filtered cells at 250g for 3 min at 4 °C. In case of a visible red pellet, aspirate the supernatant, and resuspend the pellet using 2mL of Red Blood Cell Lysis Solution (E238010) to lyse the erythrocytes at room temperature for 1 min and centrifuge at 250g for 3 min at 4°C.

8.      Aspirate the supernatant and resuspend the pellet in Cancer Organoid Basal Medium, centrifuge at 250g for 3 min at 4°C, repeat this step once more time.

9.      Aspirate the supernatant and resuspend the pellet in Matrigel. The Matrigel should be kept on ice to prevent it from solidifying. Perform the process as quickly as possible. The amount of Matrigel used depends on the size of the pellet. Approximately 10,000 cells should be plated in 25 μL of Matrigel.

CRITICAL: Do not overly dilute the Matrigel (Matrigel should be >70% (Matrigel vol/Total vol)), as this may inhibit the proper formation of the solid droplets.

10.   Plate the Matrigel containing organoids at the bottom of 24-well cell culture plates in droplets of ~30 μL each around the center of the well..

CRITICAL: Once the organoids are resuspended in Matrigel, proceed with plating as quickly as possible, as the Matrigel may solidify in the tube or pipette tip. Do not let the Matrigel touch the wall of wells.

11.   Place the culture plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15-25 min to let the Matrigel solidify.

12.   Prepare the required amount of gastric cancer organoid complete medium.

13.   Once the Matrigel droplets have solidified (15-25 min), open the plate and carefully add 500 μL of organoid complete medium to each well.

CRITICAL: Do not add the medium directly on top of the Matrigel droplets, as this might disrupt the droplets.

14.   Place the culture plate in a humidified incubator at 37 °C and 5% (vol/vol) CO2.

15.   Change the medium every 3-4 d by carefully aspirating the medium from the wells and replacing it with a fresh, pre-warmed organoid complete medium.

16.   Closely monitor the organoid formation. Ideally, patient-derived gastric cancer organoids should be passaged for the first time between 7 and 10 d after the initial plating.

Splitting and Passaging of Organoids

17.   Pipette up and down to disrupt the Matrigel and transfer the organoid suspension to a 1.5 mL conical tube.

18.   Pipette the organoid suspension up and down to mix thoroughly by pipetting against the bottom of the tube to create pressure, which will aid the removal of Matrigel.

19.   Centrifuge organoids at 250g for 3 min at room temperature.

20.   Aspirate the supernatant and split organoids using either Organoid Dissociation Solution (E238001) or by mechanical disruption. For Organoid Dissociation Solution-based cell dissociation, resuspend the pellet in 0.2 mL of Organoid Dissociation Solution, pipette up and down and incubate at 37 °C until organoids fall apart. Pipette up and down with a filter tip for ≥8 times every 2 min to aid in the disruption of the organoids. Closely monitor the digestion to keep the incubation time in the Organoid Dissociation Solution to a minimum. In case of mechanical disruption, resuspend the pellet in 1.5 mL of Cancer Organoid Basal Medium. Carefully pipette the organoid suspension up and down 30 times by pipetting against the bottom of the tube to create pressure, which will aid organoid disruption.

CRITICAL: Do not dissociate in Organoid Dissociation Solution for >5 min, as this may result in poor organoid outgrowth or even loss of the culture. As a rule of thumb, digestion is complete if a mixture of small lumps of cells (consisting of 10–50 cells) can be observed.

21.   After shearing is complete, wash once by topping up with 1 mL of Cancer Organoid Basal Medium, and centrifuge at 250g for 3 min at room temperature.

22.   Aspirate the supernatant and resuspend the organoid pellet in 70% (vol/vol) Matrigel, and plate organoids in droplets at the bottom of a culture plate as described in Steps 10. After plating is complete, transfer the plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15–25 min.

23.   Pre-warm gastric cancer organoid complete medium at 37 °C.

24.   After the Matrigel droplets have solidified (15–25 min), carefully pipette pre-warmed medium into the wells.

25.   Place culture plates in a humidified incubator at 37 °C and 5% (vol/vol) CO2 until the organoids are needed for further experiments.

bioGenous小细胞肺癌 Small Cell Lung Cancer Organoid Kit(K2121-SL)

小细胞肺癌 Small Cell Lung Cancer Organoid Kit

货号:K2121-SL

规格:100ml

500ml

品牌:bioGenous

产品介绍

Product Description:

bioGenousTM Small Cell Lung Cancer Organoid Kit is a chemically defined cell culture medium for the establishment and maintenance of human Small Cell Lung Cancer organoids. Patient-derived cancer organoids recapitulate the genomic and pathological features of original tumors and therefore hold great promise for medical research and precision medicine.


技术参数

Product Information:

Component

Component Cat#

Volume

Storage & Stability

bioGenousTM Small Cell Lung Cancer Organoid Basal Medium

K2121-SL-A100/A500

100mL/500mL

4℃,12 months

bioGenousTM Small Cell Lung Cancer Organoid Supplement B (50x)

K2121-SL-B100/B500

2mL/10mL

-20, avoid repeated freeze-thaw cycles, 12 months

bioGenousTM Small Cell Lung Cancer Organoid Supplement C (250x)

K2121-SL-C100/C500

0.4mL/2mL

-20, avoid repeated freeze-thaw cycles, 12 months

Preparation of Small Cell Lung Cancer Organoid Complete Medium

Use a sterile technique to prepare the small cell lung cancer organoid complete medium. The following example is for preparing 10mL complete medium. If preparing other volumes, adjust accordingly.

1.      Thaw Small Cell Lung Cancer Organoid Supplement B(50x) and Small Cell Lung Cancer Organoid Supplement C(250x) on ice. Mix thoroughly.

NOTE: Once thawed, use immediately or aliquot and store at -20°C for not more than 10 months. After thawing the aliquots, use immediately. Do not re-freeze.

2.      Add 200μL Small Cell Lung Cancer Organoid Supplement B(50x) and 40μL Small Cell Lung Cancer Organoid Supplement C(250x) to 9.76mL Small Cell Lung Cancer Organoid Basal Medium. Mix thoroughly.

NOTE: If not used immediately, store the complete medium at 2-8°C for not more than 2 weeks. The Small Cell Lung Cancer Organoid Supplement B contains fungicide and antibiotics (50x).

Protocol for Establishment of Patient-Derived Small Cell Lung Cancer Organoids

CAUTION Studies involving primary human tissue material must follow all relevant institutional and government regulations. Informed consent must be obtained from all subjects before the collection of the primary human tissue material.

Establishment of Organoids from Primary Tissue

1.      Collect primary human small cell lung cancer tissue pieces in ice-cold Primary Tissue Storage Solution (K601005) with conical tubes. Keep tissue samples at 4°C until the start of the isolation.

2.      Assess whether the obtained tissue pieces consist purely of epithelium. If fat or muscle tissues are present, remove these non-epithelial components as much as possible using surgical scissors or scalpels and forceps under a dissection microscope. If no fat or muscle tissues are present, continue to the next step immediately.

3.      Rinse the tissue with Cancer Organoid Basal Medium (B213152) or DPBS twice.

4.      Mince the tissue into small fragments of 1-3 mm3 in a cell culture dish using surgical scissors or scalpels.

5.      Digest the tissue fragments with 10mL of Tumor Tissue Digestion Solution (K601003) in a 15mL conical tube at 37°C, with variable incubation times ranging from 30 min to 1 h. Carefully monitor the digestion process, mixing the content of the tube every 5-10 min by shaking vigorously or pipetting the mixture up and down. The digestion process could be finished when most of tissue fragments are able to pass through the 1mL pipette tips.

6.      Add FBS to the tissue digestion mixture to a final concentration of 2%, and filter using a 100 μm cell strainer.

7.      Collect and centrifuge the filtered cells at 250g for 3 min at 4 °C. In case of a visible red pellet, aspirate the supernatant, and resuspend the pellet using 2mL of Red Blood Cell Lysis Solution (E238010) to lyse the erythrocytes at room temperature for 1 min and centrifuge at 250g for 3 min at 4°C.

8.      Aspirate the supernatant and resuspend the pellet in Cancer Organoid Basal Medium, centrifuge at 250g for 3 min at 4°C, repeat this step once more time.

9.      Aspirate the supernatant and resuspend the pellet in Matrigel. The Matrigel should be kept on ice to prevent it from solidifying. Perform the process as quickly as possible. The amount of Matrigel used depends on the size of the pellet. Approximately 10,000 cells should be plated in 25 μL of Matrigel.

CRITICAL: Do not overly dilute the Matrigel (Matrigel should be >70% (Matrigel vol/Total vol)), as this may inhibit the proper formation of the solid droplets.

10.   Plate the Matrigel containing organoids at the bottom of 24-well cell culture plates in droplets of ~30 μL each around the center of the well..

CRITICAL: Once the organoids are resuspended in Matrigel, proceed with plating as quickly as possible, as the Matrigel may solidify in the tube or pipette tip. Do not let the Matrigel touch the wall of wells.

11.   Place the culture plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15-25 min to let the Matrigel solidify.

12.   Prepare the required amount of small cell lung cancer organoid complete medium.

13.   Once the Matrigel droplets have solidified (15-25 min), open the plate and carefully add 500 μL of organoid complete medium to each well.

CRITICAL: Do not add the medium directly on top of the Matrigel droplets, as this might disrupt the droplets.

14.   Place the culture plate in a humidified incubator at 37 °C and 5% (vol/vol) CO2.

15.   Change the medium every 3-4 d by carefully aspirating the medium from the wells and replacing it with a fresh, pre-warmed organoid complete medium.

16.   Closely monitor the organoid formation. Ideally, patient-derived small cell lung cancer organoids should be passaged for the first time between 7 and 10 d after the initial plating.

Splitting and Passaging of Organoids

17.   Pipette up and down to disrupt the Matrigel and transfer the organoid suspension to a 1.5 mL conical tube.

18.   Pipette the organoid suspension up and down to mix thoroughly by pipetting against the bottom of the tube to create pressure, which will aid the removal of Matrigel.

19.   Centrifuge organoids at 250g for 3 min at room temperature.

20.   Aspirate the supernatant and split organoids using either Organoid Dissociation Solution (E238001) or by mechanical disruption. For Organoid Dissociation Solution-based cell dissociation, resuspend the pellet in 0.2 mL of Organoid Dissociation Solution, pipette up and down and incubate at 37 °C until organoids fall apart. Pipette up and down with a filter tip for ≥3 times every 1 min to aid in the disruption of the organoids. Closely monitor the digestion to keep the incubation time in the Organoid Dissociation Solution to a minimum. In case of mechanical disruption, resuspend the pellet in 1.5 mL of Cancer Organoid Basal Medium. Carefully pipette the organoid suspension up and down 30 times by pipetting against the bottom of the tube to create pressure, which will aid organoid disruption.

CRITICAL: Do not dissociate in Organoid Dissociation Solution for >3 min, as this may result in poor organoid outgrowth or even loss of the culture. As a rule of thumb, digestion is complete if a mixture of small lumps of cells (consisting of 10–50 cells) can be observed.

21.   After shearing is complete, wash once by topping up with 1 mL of Cancer Organoid Basal Medium, and centrifuge at 250g for 3 min at room temperature.

22.   Aspirate the supernatant and resuspend the organoid pellet in 70% (vol/vol) Matrigel, and plate organoids in droplets at the bottom of a culture plate as described in Steps 10. After plating is complete, transfer the plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15–25 min.

23.   Pre-warm small cell lung cancer organoid complete medium at 37 °C.

24.   After the Matrigel droplets have solidified (15–25 min), carefully pipette pre-warmed medium into the wells.

25.   Place culture plates in a humidified incubator at 37 °C and 5% (vol/vol) CO2 until the organoids are needed for further experiments.

bioGenous胰腺癌 Pancreatic Cancer Organoid Kit(K2101-PC)

胰腺癌 Pancreatic Cancer Organoid Kit

货号:K2101-PC

规格:100ml

500ml

品牌:bioGenous

产品介绍

Product Description:

bioGenousTM Pancreatic Cancer Organoid Kit is a chemically defined cell culture medium for the establishment and maintenance of human pancreatic cancer organoids. Patient-derived cancer organoids recapitulate the genomic and pathological features of original tumors and therefore hold great promise for medical research and precision medicine.


技术参数

Product Information:

Component

Component Cat#

Volume

Storage & Stability

bioGenousTM Pancreatic Cancer Organoid Basal Medium

K2101-PC-A100/A500

100mL/500mL

4℃,12 months

bioGenousTM Pancreatic Cancer Organoid Supplement B (50x)

K2101-PC-B100/B500

2mL/10mL

-20, avoid repeated freeze-thaw cycles, 12 months

bioGenousTM Pancreatic Cancer Organoid Supplement C (250x)

K2101-PC-C100/C500

0.4mL/2mL

-20, avoid repeated freeze-thaw cycles, 12 months

Preparation of Pancreatic Cancer Organoid Complete Medium

Use a sterile technique to prepare the pancreatic cancer organoid complete medium. The following example is for preparing 10mL complete medium. If preparing other volumes, adjust accordingly.

1.      Thaw Pancreatic Cancer Organoid Supplement B(50x) and Pancreatic Cancer Organoid Supplement C(250x) on ice. Mix thoroughly.

NOTE: Once thawed, use immediately or aliquot and store at -20°C for not more than 10 months. After thawing the aliquots, use immediately. Do not re-freeze.

2.      Add 200μL Pancreatic Cancer Organoid Supplement B(50x) and 40μL Pancreatic Cancer Organoid Supplement C(250x) to 9.76mL Pancreatic Cancer Organoid Basal Medium. Mix thoroughly.

NOTE: If not used immediately, store the complete medium at 2-8°C for not more than 2 weeks. The Pancreatic Cancer Organoid Supplement B contains fungicide and antibiotics (50x).

Protocol for Establishment of Patient-Derived Pancreatic Cancer Organoids

CAUTION Studies involving primary human tissue material must follow all relevant institutional and government regulations. Informed consent must be obtained from all subjects before the collection of the primary human tissue material.

Establishment of Organoids from Primary Tissue

1.      Collect primary human pancreatic cancer tissue pieces in ice-cold Primary Tissue Storage Solution (K601005) with conical tubes. Keep tissue samples at 4°C until the start of the isolation.

2.      Assess whether the obtained tissue pieces consist purely of epithelium. If fat or muscle tissues are present, remove these non-epithelial components as much as possible using surgical scissors or scalpels and forceps under a dissection microscope. If no fat or muscle tissues are present, continue to the next step immediately.

3.      Rinse the tissue with Cancer Organoid Basal Medium (B213152) or DPBS twice.

4.      Mince the tissue into small fragments of 1-3 mm3 in a cell culture dish using surgical scissors or scalpels.

5.      Digest the tissue fragments with 10mL of Tumor Tissue Digestion Solution (K601003) in a 15mL conical tube at 37°C, with variable incubation times ranging from 30 min to 1.5 h. Carefully monitor the digestion process, mixing the content of the tube every 5-10 min by shaking vigorously or pipetting the mixture up and down. The digestion process could be finished when most of tissue fragments are able to pass through the 1mL pipette tips.

6.      Add FBS to the tissue digestion mixture to a final concentration of 2%, and filter using a 100 μm cell strainer.

7.      Collect and centrifuge the filtered cells at 250g for 3 min at 4 °C. In case of a visible red pellet, aspirate the supernatant, and resuspend the pellet using 2mL of Red Blood Cell Lysis Solution (E238010) to lyse the erythrocytes at room temperature for 1 min and centrifuge at 250g for 3 min at 4°C.

8.      Aspirate the supernatant and resuspend the pellet in Cancer Organoid Basal Medium, centrifuge at 250g for 3 min at 4°C, repeat this step once more time.

9.      Aspirate the supernatant and resuspend the pellet in Matrigel. The Matrigel should be kept on ice to prevent it from solidifying. Perform the process as quickly as possible. The amount of Matrigel used depends on the size of the pellet. Approximately 10,000 cells should be plated in 25 μL of Matrigel.

CRITICAL: Do not overly dilute the Matrigel (Matrigel should be >70% (Matrigel vol/Total vol)), as this may inhibit the proper formation of the solid droplets.

10.   Plate the Matrigel containing organoids at the bottom of 24-well cell culture plates in droplets of ~30 μL each around the center of the well..

CRITICAL: Once the organoids are resuspended in Matrigel, proceed with plating as quickly as possible, as the Matrigel may solidify in the tube or pipette tip. Do not let the Matrigel touch the wall of wells.

11.   Place the culture plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15-25 min to let the Matrigel solidify.

12.   Prepare the required amount of pancreatic cancer organoid complete medium.

13.   Once the Matrigel droplets have solidified (15-25 min), open the plate and carefully add 500 μL of organoid complete medium to each well.

CRITICAL: Do not add the medium directly on top of the Matrigel droplets, as this might disrupt the droplets.

14.   Place the culture plate in a humidified incubator at 37 °C and 5% (vol/vol) CO2.

15.   Change the medium every 3-4 d by carefully aspirating the medium from the wells and replacing it with a fresh, pre-warmed organoid complete medium.

16.   Closely monitor the organoid formation. Ideally, patient-derived pancreatic cancer organoids should be passaged for the first time between 7 and 10 d after the initial plating.

Splitting and Passaging of Organoids

17.   Pipette up and down to disrupt the Matrigel and transfer the organoid suspension to a 1.5 mL conical tube.

18.   Pipette the organoid suspension up and down to mix thoroughly by pipetting against the bottom of the tube to create pressure, which will aid the removal of Matrigel.

19.   Centrifuge organoids at 250g for 3 min at room temperature.

20.   Aspirate the supernatant and split organoids using either Organoid Dissociation Solution (E238001) or by mechanical disruption. For Organoid Dissociation Solution-based cell dissociation, resuspend the pellet in 0.2 mL of Organoid Dissociation Solution, pipette up and down and incubate at 37 °C until organoids fall apart. Pipette up and down with a filter tip for ≥8 times every 2 min to aid in the disruption of the organoids. Closely monitor the digestion to keep the incubation time in the Organoid Dissociation Solution to a minimum. In case of mechanical disruption, resuspend the pellet in 1.5 mL of Cancer Organoid Basal Medium. Carefully pipette the organoid suspension up and down 30 times by pipetting against the bottom of the tube to create pressure, which will aid organoid disruption.

CRITICAL: Do not dissociate in Organoid Dissociation Solution for >7 min, as this may result in poor organoid outgrowth or even loss of the culture. As a rule of thumb, digestion is complete if a mixture of small lumps of cells (consisting of 10–50 cells) can be observed.

21.   After shearing is complete, wash once by topping up with 1 mL of Cancer Organoid Basal Medium, and centrifuge at 250g for 3 min at room temperature.

22.   Aspirate the supernatant and resuspend the organoid pellet in 70% (vol/vol) Matrigel, and plate organoids in droplets at the bottom of a culture plate as described in Steps 10. After plating is complete, transfer the plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15–25 min.

23.   Pre-warm pancreatic cancer organoid complete medium at 37 °C.

24.   After the Matrigel droplets have solidified (15–25 min), carefully pipette pre-warmed medium into the wells.

25.   Place culture plates in a humidified incubator at 37 °C and 5% (vol/vol) CO2 until the organoids are needed for further experiments.

bioGenous子宫内膜癌 Endometrial Cancer Organoid Kit(K2166-EC)

子宫内膜癌 Endometrial Cancer Organoid Kit

货号:K2166-EC

规格:100ml

500ml

品牌:bioGenous

产品介绍

Product Description:

bioGenousTM Endometrial Cancer Organoid Kit is a chemically defined cell culture medium for the establishment and maintenance of human endometrial cancer organoids. Patient-derived cancer organoids recapitulate the genomic and pathological features of original tumors and therefore hold great promise for medical research and precision medicine.


技术参数

Product Information:

Component

Component Cat#

Volume

Storage & Stability

bioGenousTM Endometrial Cancer Organoid Basal Medium

K2166-EC-A100/A500

100mL/500mL

4℃,12 months

bioGenousTM Endometrial Cancer Organoid Supplement B (50x)

K2166-EC-B100/B500

2mL/10mL

-20, avoid repeated freeze-thaw cycles, 12 months

bioGenousTM Endometrial Cancer Organoid Supplement C (250x)

K2166-EC-C100/C500

0.4mL/2mL

-20, avoid repeated freeze-thaw cycles, 12 months

Preparation of Endometrial Cancer Organoid Complete Medium

Use a sterile technique to prepare the endometrial cancer organoid complete medium. The following example is for preparing 10mL complete medium. If preparing other volumes, adjust accordingly.

1.      Thaw Endometrial Cancer Organoid Supplement B(50x) and Endometrial Cancer Organoid Supplement C(250x) on ice. Mix thoroughly.

NOTE: Once thawed, use immediately or aliquot and store at -20°C for not more than 10 months. After thawing the aliquots, use immediately. Do not re-freeze.

2.      Add 200μL Endometrial Cancer Organoid Supplement B(50x) and 40μL Endometrial Cancer Organoid Supplement C(250x) to 9.76mL Endometrial Cancer Organoid Basal Medium. Mix thoroughly.

NOTE: If not used immediately, store the complete medium at 2-8°C for not more than 2 weeks. The Endometrial Cancer Organoid Supplement B contains fungicide and antibiotics (50x).

Protocol for Establishment of Patient-Derived Endometrial Cancer Organoids

CAUTION Studies involving primary human tissue material must follow all relevant institutional and government regulations. Informed consent must be obtained from all subjects before the collection of the primary human tissue material.

Establishment of Organoids from Primary Tissue

1.      Collect primary human endometrial cancer tissue pieces in ice-cold Primary Tissue Storage Solution (K601005) with conical tubes. Keep tissue samples at 4°C until the start of the isolation.

2.      Assess whether the obtained tissue pieces consist purely of epithelium. If fat or muscle tissues are present, remove these non-epithelial components as much as possible using surgical scissors or scalpels and forceps under a dissection microscope. If no fat or muscle tissues are present, continue to the next step immediately.

3.      Rinse the tissue with Cancer Organoid Basal Medium (B213152) or DPBS twice.

4.      Mince the tissue into small fragments of 1-3 mm3 in a cell culture dish using surgical scissors or scalpels.

5.      Digest the tissue fragments with 10mL of Tumor Tissue Digestion Solution (K601003) in a 15mL conical tube at 37°C, with variable incubation times ranging from 30 min to 1 h. Carefully monitor the digestion process, mixing the content of the tube every 5-10 min by shaking vigorously or pipetting the mixture up and down. The digestion process could be finished when most of tissue fragments are able to pass through the 1mL pipette tips.

6.      Add FBS to the tissue digestion mixture to a final concentration of 2%, and filter using a 100 μm cell strainer.

7.      Collect and centrifuge the filtered cells at 250g for 3 min at 4 °C. In case of a visible red pellet, aspirate the supernatant, and resuspend the pellet using 2mL of Red Blood Cell Lysis Solution (E238010) to lyse the erythrocytes at room temperature for 1 min and centrifuge at 250g for 3 min at 4°C.

8.      Aspirate the supernatant and resuspend the pellet in Cancer Organoid Basal Medium, centrifuge at 250g for 3 min at 4°C, repeat this step once more time.

9.      Aspirate the supernatant and resuspend the pellet in Matrigel. The Matrigel should be kept on ice to prevent it from solidifying. Perform the process as quickly as possible. The amount of Matrigel used depends on the size of the pellet. Approximately 10,000 cells should be plated in 25 μL of Matrigel.

CRITICAL: Do not overly dilute the Matrigel (Matrigel should be >70% (Matrigel vol/Total vol)), as this may inhibit the proper formation of the solid droplets.

10.   Plate the Matrigel containing organoids at the bottom of 24-well cell culture plates in droplets of ~30 μL each around the center of the well..

CRITICAL: Once the organoids are resuspended in Matrigel, proceed with plating as quickly as possible, as the Matrigel may solidify in the tube or pipette tip. Do not let the Matrigel touch the wall of wells.

11.   Place the culture plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15-25 min to let the Matrigel solidify.

12.   Prepare the required amount of endometrial cancer organoid complete medium.

13.   Once the Matrigel droplets have solidified (15-25 min), open the plate and carefully add 500 μL of organoid complete medium to each well.

CRITICAL: Do not add the medium directly on top of the Matrigel droplets, as this might disrupt the droplets.

14.   Place the culture plate in a humidified incubator at 37 °C and 5% (vol/vol) CO2.

15.   Change the medium every 3-4 d by carefully aspirating the medium from the wells and replacing it with a fresh, pre-warmed organoid complete medium.

16.   Closely monitor the organoid formation. Ideally, patient-derived endometrial cancer organoids should be passaged for the first time between 7 and 10 d after the initial plating.

Splitting and Passaging of Organoids

17.   Pipette up and down to disrupt the Matrigel and transfer the organoid suspension to a 1.5 mL conical tube.

18.   Pipette the organoid suspension up and down to mix thoroughly by pipetting against the bottom of the tube to create pressure, which will aid the removal of Matrigel.

19.   Centrifuge organoids at 250g for 3 min at room temperature.

20.   Aspirate the supernatant and split organoids using either Organoid Dissociation Solution (E238001) or by mechanical disruption. For Organoid Dissociation Solution-based cell dissociation, resuspend the pellet in 0.2 mL of Organoid Dissociation Solution, pipette up and down and incubate at 37 °C until organoids fall apart. Pipette up and down with a filter tip for ≥8 times every 2 min to aid in the disruption of the organoids. Closely monitor the digestion to keep the incubation time in the Organoid Dissociation Solution to a minimum. In case of mechanical disruption, resuspend the pellet in 1.5 mL of Cancer Organoid Basal Medium. Carefully pipette the organoid suspension up and down 30 times by pipetting against the bottom of the tube to create pressure, which will aid organoid disruption.

CRITICAL: Do not dissociate in Organoid Dissociation Solution for >7 min, as this may result in poor organoid outgrowth or even loss of the culture. As a rule of thumb, digestion is complete if a mixture of small lumps of cells (consisting of 10–50 cells) can be observed.

21.   After shearing is complete, wash once by topping up with 1 mL of Cancer Organoid Basal Medium, and centrifuge at 250g for 3 min at room temperature.

22.   Aspirate the supernatant and resuspend the organoid pellet in 70% (vol/vol) Matrigel, and plate organoids in droplets at the bottom of a culture plate as described in Steps 10. After plating is complete, transfer the plate into a humidified incubator at 37 °C and 5% (vol/vol) CO2 for 15–25 min.

23.   Pre-warm endometrial cancer organoid complete medium at 37 °C.

24.   After the Matrigel droplets have solidified (15–25 min), carefully pipette pre-warmed medium into the wells.

Place culture plates in a humidified incubator at 37 °C and 5% (vol/vol) CO2 until the organoids are needed for further experiments.