Скачать с ютуб Sample Preparation Tutorial Part II: How to Cut Your Frozen Tissue Sample To Get Beautiful Sections? в хорошем качестве

Sample Preparation Tutorial Part II: How to Cut Your Frozen Tissue Sample To Get Beautiful Sections?

Are you having trouble sectioning your tissue sample for the Molecular Cartography workflow? Here are the tips and tricks to make beautiful sections and basic troubleshooting. For more technical questions, please get in touch with our regional Customer Technology Advisor or send us an email at [email protected]. We are always happy to help if you have any issues during tissue preparation. Full transcript: Hello, my name is Jeroen, and I'm a Customer Technology Advisor with Resolve Biosciences. Today, I will be showing you how to cut your tissue sample and transfer those sections onto Molecular Cartography glass slides. I will be cutting 10-micrometer sections on Leica cryostat. For detailed instructions on how to use your cryostat, please consult your device manual. Carefully remove the provided Molecular Cartography slide from the cartridge box on dry ice. Note that marked captured areas are on the underside of the slide. Place the slide, the frozen embedded sample, brushes, forceps, and scalpel in the cryostat and wait for at least 30 minutes to allow everything to equilibrate to cryostat temperature. Don't forget to clean the knife with ethanol before proceeding. Mount the sample on the specimen disc with OCT and position it in the cryostat object holder. First, I usually apply a base of OCT for better adhesion. Make sure the glass slide is on a flat surface in a cryostat chamber before transferring the sections. Move the sample close to the knife to start cutting. For Molecular Cartography, we need 10 micrometer thick sections. Adjust the anti-roll guide to ensure sections are flat. Alternatively, you can use a brush to guide the section over the carrier to flatten it. Avoid overstretching or tearing your sections. If the sample is too big to fit into the capture area, cut out the desired area of the tissue with a scalpel. Alternatively, you can trim the tissue block to the appropriate size using the scalpel. When doing so, make sure to retract the tissue slightly before making a new section to prevent tissue distortion. In order not to break the glass slide, use toothbrushes to place the section in the center of the marked area of the slide. Ensure that no part of the sample or OCT is positioned outside of the marked area. If samples are not positioned correctly, the wells cannot be sealed and we will be unable to analyze the samples. Gently flattened sections to ensure good surface contact. Slightly warm sections to adhere them to the slide. We use old-fashioned body heat in the form of your finger underneath the slides. Do not apply pressure to prevent the glass slide from breaking and make sure the edges of the sections are well heated to prevent detachment. Always do this procedure inside the cryostat chamber to prevent condensation on the glass slide. Place the slide on the surface of the cryostat for the sections to freeze. Continue repeating this process until all eight samples have been transferred to the slides. Place finished slides back into the provided Molecular Cartography cartridge box. Store samples at -80 degrees Celsius or on dry ice for shipping and handling. For shipping, fill a styrofoam box completely with dry ice. Once we receive the samples, we will then process them for imaging. We will attach a plastic separator to the slides. The plastic separators will create separate compartments for individual staining. If samples are not positioned correctly, the wells cannot be sealed and we will be unable to analyze the samples. Having trouble making beautiful sections? Here are some basic troubleshooting tips that can help you out when horizontal stripes appear on the section. It can be that the cryostat temperature is too cold. Adjust to a warmer temperature until the surface of the sections appears smooth. If the sections appear to be folded or have waves in them, the cryostat temperature might be too warm. Adjust to a colder temperature to make smoother sections. #spatialtranscriptomics #spatialbiology #tissuepreparation