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In this video, we demonstrate how to achieve efficient and gentle tissue dissociation using the STEMprep™ Tissue Dissociator.

STEMprep™ Tissue Dissociator automates tissue dissociation to generate high-viability single-cell suspensions for downstream applications such as cell separation, cell culture, flow cytometry, and immunology or cancer research. With built-in temperature control and modular scalability, it ensures efficient, reproducible results while preserving cell integrity. STEMprep™ Tissue Dissociator can have up to four units connected together, which allows the processing of up to 16 different samples simultaneously.

Part One: Before Tissue Dissociation.

Refer to the Product Information Sheet that comes with your tissue-specific dissociation kit for the recommended protocol. Assess your needs based on the number of samples to be processed. To begin processing tissues with the STEMprep™ Tissue Dissociator, make sure you have gathered the necessary materials, including your tissue samples, a STEMprep™ dissociation kit, STEMprep™ Sample Tubes, EasySep™ Buffer or PBS containing 2% FBS and 1 mM EDTA, 50 mL conical tubes, a plastic rack for centrifuge tubes, and 70 μm cell strainers. If desired, the STEMprep™ Tissue Dissociator can be operated in a biosafety cabinet.

Initialize the STEMprep™ Tissue Dissociator. Select a User Account from the Control Bar. User Accounts can be used to better organize custom protocols and prevent others from accidentally modifying or deleting them. User Accounts can be created and managed under the Settings menu. The Custom Protocol Editor provides granular control over all key dissociation parameters: time, temperature, and mechanical forces, speed, and direction on a step-by-step basis. For more information on creating your own tissue-specific custom protocols, please refer to the User Manual.

Select the slots to run a protocol using the Run Control screen. Multiple slots can be selected at once to run the same protocol simultaneously. Slots can also run different protocols simultaneously. In this case, we will only be running each protocol on a single slot. So, we will cancel and select a single slot and the desired protocol. The Hold Temperature function can be used to keep the sample at a specific temperature after the run completes. The screen display will vary depending on whether you have one, two, three, or four STEMprep™ units installed. In this experiment, we will run Mouse Brain, Liver, Lung, and Spleen protocols.

For recommendations on Mouse Tumor samples, please refer to the STEMprep™ Mouse Tumor Dissociation Kit’s Product Information Sheet. A Homogenization protocol is also available to facilitate molecular analysis.

Part Two: Dissociating Tissue Samples.

To maintain high cell viability, it's crucial to harvest and process mouse tissue samples immediately after sacrifice. Load diluent, enzymes, and the tissue sample into the sample tube. For the volume of diluent and tissue-specific enzymes to add, please refer to the tissue-specific Product Information Sheet. Please note that for lungs, the lobes need to be separated to ensure proper dissociation. Hard and medium solid tumor samples will also require pre-cutting for best results. Recommended tissue preprocessing steps can also be found in the tissue-specific Product Information Sheet.

Ensure the sample tube cap is properly tightened. Load the sample tube into a slot on the STEMprep™ and ensure that the tabs of the sample tube fit into the grooves of the slot. Lower the shield to the lowest position. When the shield is fully lowered, the orange bars indicating the shield is up will disappear and the run button will become active. If the sample tube is not properly loaded, the screen will display an icon indicating that the tube is not positioned correctly or that the spindle was not able to line up correctly with the rotor. Lift the shield and make sure that the tabs are properly aligned with the grooves. Then, lower the shield. If the spindle fails to engage the rotor correctly, lift the shield, turn the spindle slightly, and then lower the shield. For a complete list of different icons and what they represent, please refer to the User Manual.

Once the run begins, the remaining protocol time will be displayed. If a Hold Temperature is selected, the sample will be maintained at the specified temperature for up to 24 hours after the protocol ends. Once the protocol is completed, the dissociated sample should be a uniform single-cell suspension. We recommend filtering the dissociated sample through a 70 μm cell strainer. Pre-wet the cell strainer with buffer before filtering the sample. Here we chose EasySep™ Buffer. Rinse the sample tube and collect the remaining cells. Once finished, add buffer to the collection tube until it reaches 50 mL. Your tissue sample is now dissociated into a single-cell suspension.

Next, we will prepare the dissociated tissue for downstream applications. In a centrifuge, spin the collection tube at 300 x g for 10 minutes at room temperature. After centrifugation, dissociated brain and liver tissue may be loose. Do not decant the supernatant. Instead, carefully aspirate it. Other tissues such as spleen or lung form a more tightly packed pellet, and the supernatant can be poured off. Please refer to the tissue-specific Product Information Sheet for recommended additional cleanup steps.

Ammonium chloride lysis buffer can be used to lyse red blood cells, and OptiPrep™ Density Medium can be adjusted to remove myelin or tissue debris from samples. Depending on the tissue type, various markers can be used to assess the yield and identify the cell subsets present. It is recommended to include a viability dye in the flow cytometry analysis. STEMprep™ is also compatible with EasySep™ cell isolation kits and �����ƽ�� culture media.

Elevate your tissue dissociation workflow with STEMprep™. Learn more at stemcell.com/STEMprep.