Skin cells on slide picked up on dirty water bath
The microtomy process outlined below is useful for all types of tissue samples, but is also particularly helpful when sectioning derms. Although a wide variety of microtomy techniques exist throughout histology laboratories, the tips below are the simplest to apply and most universal.
- The flotation bath should be clean and water maintained 5-10º C below the melting point of paraffin. Higher temperatures may separate the tissue and dissolve the ribbons before they are placed on the slides. Lower temperatures will not allow the tissue to relax enough to remove the wrinkles.
- Microtome should be on a solid/stable working surface and free of vibration. All clamps should be tightened before beginning. A slow and steady rotation of the hand wheel also reduces vibration and artifacts in sections, such as undulations and venetian blinds. The top image shows this artifact.
- Tissue blocks should be coarse-faced (or surfaced) and chilled on ice prior to microtomy. Adding a small amount of water to the ice helps dry tissue soften, improving sectioning and reducing chatter. Nails can be softened in a weak ammonia solution or fabric softener prior to sectioning.
- Skin orientation should be inspected after facing the block to ensure that all of the skin samples are embedded correctly and that all of the tissue is exposed on the block surface. Melt down blocks and re-embed when necessary.
- Adhesive or charged slides should be used when sectioning calcified tissue or nails, as well as sections for special stains.
- Blocks should be secured in block holder with hair in tissue pointing up and away from the blade. For optimal sections, the blade should cut through the fat and dermis first, and the epidermis and hair last.
- Routine sections should be cut at 3-5 microns. Skin sections are optimal at 4 microns. Only one block at a time should be sectioned and floated on the water to prevent tissue transfer or mix-ups.
- Double check the block identification against the slide to ensure accuracy before placing tissue on slide.
- The floating ribbon should be gently stretched to eliminate wrinkles in the sections. Skin sections are prone to wrinkles due to the elastic fibers present, as well as the competing layers of skin.
- Surface of the flotation bath should be skimmed (with lint free paper wipes) between each block to clean debris and prevent cross-contamination. Leaning over the bath should be avoided and wearing gloves and long sleeves should be encouraged to prevent personal skin cells from falling onto the water. Water should be changed frequently, at least once per shift. The lower image depicts skin cells on a slide.
- Finished slides should be stored vertically to allow the water to drip down and away from the sections. Sections of nails should be dried over-night at room temperature or in an incubator for several hours prior to deparaffinization to encourage adhesion.
- Microtome face-plate/blade holder should be wiped clean before proceeding to the next tissue block to prevent tissue carryover.