Christensen A K
J Cell Biol. 1971 Dec;51(3):772-804. doi: 10.1083/jcb.51.3.772.
A simple method has been developed that allows frozen thin sections of fresh-frozen tissue to be cut on a virtually unmodified ultramicrotome kept at room temperature. A bowl-shaped Dewar flask with a knifeholder in its depths replaces the stage of the microtome; a bar extends down into the bowl from the microtome's cutting arm and bears the frozen tissue near its lower end. When the microtome is operated, the tissue passes a glass or diamond knife in the depths of the bowl as in normal cutting. The cutting temperature is maintained by flushing the bowl with cold nitrogen gas, and can be set anywhere from about -160 degrees C up to about -30 degrees C. The microtome is set for a cutting thickness of 540-1000 A. Sections are picked up from the dry knife edge, and are placed on membrane-coated grids, flattened with the polished end of a copper rod, and either dried in nitrogen gas or freeze-dried. Throughout the entire process the tissue is kept cold and does not come in contact with any solvent. The morphology seen in frozen thin sections of rat pancreas and liver generally resembles that in conventional preparations, although freezing damage and low contrast limit the detail that can be discerned. Among unusual findings is a frequent abundance of mitochondrial granules in material prepared by this method.
已开发出一种简单方法，可在室温下几乎未作改动的超薄切片机上切割新鲜冷冻组织的冷冻薄切片。一个碗形杜瓦瓶，其底部有一个刀架，取代了切片机的载物台；一根杆从切片机的切割臂向下延伸到碗中，并在其下端附近支撑冷冻组织。当操作切片机时，组织像在正常切割中一样在碗的深处经过玻璃或金刚石刀。通过用冷氮气冲洗碗来维持切割温度，温度可设定在约 -160℃至约 -30℃之间的任何位置。切片机设定的切割厚度为540 - 1000埃。切片从干燥的刀刃上取下，放在涂有膜的网格上，用铜棒的抛光端展平，然后在氮气中干燥或冷冻干燥。在整个过程中，组织保持低温，且不与任何溶剂接触。大鼠胰腺和肝脏冷冻薄切片中所见的形态通常与传统制剂中的相似，尽管冷冻损伤和低对比度限制了可分辨的细节。在异常发现中，通过这种方法制备的材料中线粒体颗粒常常大量存在。
