Nucleolus microscopy

The nucleolus is composed of 5 to 10 percent RNA, and the remainder of the mass is protein and DNA. In light microscopy it appears to be spherical and basophilic (Prob. 1.1). Its function is the synthesis of ribosomal RNA (Chap. 17). There may be more than one per nucleus. 

Fig. 2 Characterization of enriched nucleoli by light and electron microscopy. (A and B) Nucleoli in sucrose from two different step gradients were viewed by phase-contrast microscopy scale bar, 10 /Ltm. (C and D) Micrographs of nucleoli are shown at two magnifications. The nuclear envelope (A) and the nucleolus (A) are marked. A micrograph of the fraction immediately above the nucleoli (corresponding to lane 1.75 in Fig. 1) is also shown (E). (C) Scale bar, 1 /urn (D) scale bar, 0.5 jum. Magnification is the same in C and E. For EM, nucleoli were fixed, embedded in Spurr s resin, poststained, and viewed using standard methods, as described previously (de Beus er al 1994).

Fig. 6 Distribution of flbrillar centers in PHA-stimulated human lymphocytes at 0 (a and d), 24 (b and e), and 48 hr (c and f) as visualized by conventional electron microscopy (a-c) and by indirect immunofluorescence (d-f) using a human autoantibody to RNA polymerase I. FC, Fibrillar center, DFC, dense flbrillar component G, granular component. The resting, unstimulated lymphocyte (a and d) contains a single large FC (arrowhead in d) that increases in number and decreases in size as the nucleolus grows in size with continued stimulation, (a-c) Scale bar, 0.5 /zm. (d-f) Scale bar, 5 /i.m.

Because the component parts of the nucleolus are often interspersed, one of the best ways to identify the location of a particular nucleolar protein is to first segregate the nucleolus into its component parts and then attempt immunolocali-zation with specific antibodies by means of immunofluorescence and/or immu-noelectron microscopy (Ochs et al, 1985b). A number of drugs have typically been employed to induce just such a segregation (Simard and Bernhard, 1966 Busch and Smetana, 1970 Abelson and Penman, 1975). Each of these gives a somewhat different (and distinctive) type of segregated nucleolus (see Table II and Fig. 7). 

The picture obtained by electron microscopy is static, whereas the real mitochondrion is a dynamic body that bends or straightens its structure and can be displaced throughout the cytoplasmic field. Motion pictures of mitochondria have even demonstrated mitochondrial fragmentation in both long and short axes and fusion of small mitochondria to form larger mitochondria. Mitochondria can travel from the cellular to the nuclear membrane where they appear to come in close contact with the nucleolus. The cause and the significance of these movements remain unknown. 

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