Abstract
The present paper reviews recent results obtained by different molecular biology-based, immunocytological approaches to the localization and identification of nucleic acids in sections of biological material. Examples of sensitive, high-resolution detection methods for RNA, DNA or specialized DNA regions are presented. Special emphasis is placed on the potential values and limitations of these new methods.
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Bendayan M (1981a) Electron microscopical localization of nucleic acids by means of nuclease-gold complexes. Histochem J 13:699–710
Bendayan M (1981b) Ultrastructural localization of nucleic acids by the use of enzyme-gold complexes. J Histochem Cytochem 29:531–541
Bendayan M (1984) Enzyme-gold electron microscopic cytochemistry: a new affinity approach for the ultrastructural localization of macromolecules. J Electron Microsc Techn 1:349–372
Bendayan M, Puvion E (1983) Ultrastructural detection of RNA: complementarity of high-resolution autoradiography and of RNase-gold method. J Ultrastruct Res 83:274–283
Bendayan M, Stephens H (1984) Double labelling cytochemistry applying the protein A-gold technique. In: Polak I Varndell I (eds) Immunolabelling of electron microscopy. Elsevier, Amsterdam, pp 143–154
Bendayan M, Nanci A, Kan F (1987) Effect of tissue processing on colloidal gold cytochemistry. J Histochem Cytochem 35:983–996
Binder M, Tourmente S, Roth J, Renaud M, Gehring W (1986) In situ hybridization at the electron microscope level: localization of transcripts on ultrathin sections of Lowicryl K4M-embedded tissue using biotinylated probes and protein-A-gold complexes. J Cell Biol 102:1646–1653
Bollum FJ (1963) Intermediate states in enzymatic DNA synthesis. J Cell Comp Physiol 62:61–71
Bollum FJ (1974) Terminal deoxynucleotidyl transferase. In: Boyer PD (ed) The enzymes, vol 10. Academic Press, New York, pp 145–171
Bouteille M, Dupuy-Coin AM, Moyne G (1975) Methods for localization of proteins and nucleoproteins in the cell nucleus by high resolution autoradiography and cytochemistry. Methods Enzymol 40:3–41
Cedar H (1975) Transcription of DNA and chromatin with calf thymus RNA polymerase B in vitro. J Mol Biol 95:257–269
Chang LMS, Bollum FJ (1986) Molecular biology of terminal transferase. Crit Rev Biochem 21:27–52
Cheniclet C, Bendayan M (1990) Comparative pyrimidine- and purine-specific RNase-gold labeling on pancreatic acinar cells and isolated hepatocytes. J Histochem Cytochem 38:551–562
Cheniclet C, Garzon S, Bendayan M (1995) In situ detection of nucleic acids by the nuclease-gold method. In: Morel G (ed) Visualization of nucleic acids. CRC Press, Boca Raton, pp 95–109
Craigh SW, Pollard TD (1982) Actin-binding proteins. Trends Biochem Sci 7:88–92
Dyer KA, Riley D, Gartler SM (1985) Analysis of inactive X chromosome structure by in situ nick-translation. Chromosoma (Berl) 92:209–213
Fakan S (1976) High-resolution autoradiography as a tool for the localization of nucleic acid synthesis and distribution in the mammalian cell nucleolus. J Microsc 106:159–171
Fakan S (1978) High resolution autoradiography studies on chromatin functions. In: Busch H (ed) The cell nucleus, vol 5, Academic Press, New York, pp 3–53
Fakan S (1986) Structural support for RNA synthesis in the cell nucleus. Methods Achiev Exp Pathol 12:105–140
Fakan S, Modak S (1973) Localization of DNA in ultrathin tissue sections incubated with terminal deoxynucleotidyl transferase, as visualized by electron microscope autoradiography. Exp Cell Res 77:95–104
Fakan S, Leser G, Martin T (1984) Ultrastructural distribution of nuclear ribonucleoproteins as visualized by immunocytochemistry on thin sections. J Cell Biol 98:358–363
Ferrarro M, Prantera G (1988) Human NORs show correlation between transcriptional activity, DNase I sensitivity and hypomethylation. Cytogenet Cell Genet 47:58–61
Gazit B, Cedar H, Lerer I, Voss R (1982) Active genes are sensitive to deoxyribonuclease I during metaphase. Science 217:648–650
Geuskens M (1977) RNA synthesis initiated in ultrathin sections of plastic-embedded cells and tissues by an exogeneous RNA polymerase. Biol Cell 28:207–214
Geuskens M, Recondo A de, Chevaillier P (1975) Autoradiographic demonstration of DNA replication in ultrathin sections of plastic-embedded tissues using an exogeneous DNA polymerase. Chromosoma 52:175–188
Graaf A de, Van Hemert F, Linnemans WAM, Brakenhoff GJ, Jong L de, Van Renswoude J, Van Driel R (1990) Three-dimensional distribution of DNase I-sensitive chromatin regions in interphase nuclei of embryonal carcinoma cells. Eur J Cell Biol 52:135–141
Gross D, Garrard W (1988) Nuclease hypersensitive sites in chromatin. Annu Rev Biochem 57:159–197
Hadjiolov A (1985) The nucleolus and ribosome biogenesis (Cell biology monographs, vol 12) Springer, Vienna New York, pp 1–263
Hutchison N, Weintraub H (1985) Localization of DNase I-sensitive sequences to specific regions of interphase nuclei. Cell 43:471–482
Hutchison N, Langer-Safer P, Ward D, Hamkalo B (1982) In situ hybridization at the electron microscope level: hybrid detection by autoradiography and colloidal gold. J Cell Biol 95:609–618
Jacob J, Todd K, Birnstiel M, Bird A (1971) Molecular hybridization of3H-labelled ribosomal RNA with DNA in ultrathin sections prepared for electron microscopy. Biochem Biophys Acta 228:761–766
Kelly R, Cozzarelli NR, Deutscher MP, Lehman IR, Kornberg A (1970) Enzymatic synthesis of deoxyribonucleic acid. XXXII. Replication of duplex deoxyribonucleic acid by polymerase at a single strand break. J Biol Chem 245:39–45
Kerem BS, Goitein R, Richler C, Marcus M, Cedar H (1983) In situ nick-translation distinguishes between active and inactive X chromosomes. Nature 304:88–90
Kerem BS, Goitein R, Diamond G, Cedar H, Marcus M (1984) Mapping of DNase I sensitive regions on mitotic chromosomes. Cell 38:493–499
Knezetic J, Luse D (1986) The presence of nucleosomes on a DNA template prevents initiation by RNA polymerase II in vitro. Cell 45:95–104
Krystosek A, Puck TT (1990) The spatial distribution of exposed nuclear DNA in normal, cancer, and reverse-transformed cells. Proc Natl Acad Sci USA 87:6560–6564
Kuo M, Plunkett W (1985) Nick-translation of metaphase chromosomes. In vitro labeling of nuclease-hypersensitive regions in chromosomes. Proc. Natl Acad Sci USA 82:854–858
Levitt A, Axel R, Cedar H (1979) Nick-translation of active genes in intact nuclei. Dev Biol 69:496–505
Manuclidis L, Langer-Safer PR, Ward DC (1982) High resolution mapping of satellite DNA using biotin-labeled DNA probes. J Cell Biol 95:619–625
McGhee JD, Felsenfeld G (1980) Nucleosome structure. Annu Rev Biochem 49:1115–1156
Meilhac M, Chambon P (1973) Animal DNA-dependent RNA polymerases. Initiation sites on calf-thymus DNA. Eur J Biochem 35:454–463
Modak SP, Bollum FJ (1970) Terminal lens cell differentiation. III. Initiator activity of DNA during nuclear degeneration. Exp Cell Res 62:421–432
Modak SP, Donnelly GM, Karasaki S, Hardin CV, Reddan J, Unakar N (1973) An enzymatic assay for DNA in epoxy-tissue sections by light microscope-autoradiography. Exp Cell Res 76:218–222
Morel G, Le Guellec D, Mertani H, Tremblean A (1995) In situ hybridization for electron microscopy. In: Morel G (ed) Visualization of nucleic acids. CRC Press, Boca Raton, pp 229–257
Morse R, Simpson R (1988) DNA in the nucleosome. Cell 54:285–287
Murer-Orlando M, Peterson A (1985) In situ nick-translation of human and mouse chromosomes detected with a biotinylated nucleotide. Exp Cell Res 157:322–334
Olmedilla A, Testillano P, Raska I, Risueno MC (1992) In situ nick-translation and anti-BrdU techniques as convenient tools to study the functional regions of chromatin in plants. Electron Microscopy EUREM 92, Granada, Spain, part 3, pp 193–194
Reeves R (1984) Transcriptionally active chromatin. Biochem Biophys Acta 782:348–393
Scheer U, Benavente R (1990) Functional and dynamic aspects of the mammalian nucleolus. Bioessays 12:14–21
Scheer U, Rose K (1984) Localization or RNA polymerase I in interphase cells and mitotic chromosomes by light and electron microscopic immunocytochemistry. Proc Natl Acad Sci USA 81:1431–1435
Scheer U, Thiry M, Goessens G (1993) Structure, function and assembly of the nucleolus. Trends Cell Biol 3:236–241
Sippel AE (1973) Purification and characterization of adenosine triphosphate: ribonucleic acid adenyltransferase fromEscherichia coli. Eur J Biochem 37:31–40
Spector D, Fu XD, Maniatis T (1991) Associations between distinct pre-mRNA splicing components and the cell nucleus. EMBO J 10:3467–3481
Sperling K, Kerem BS, Goitein R, Kottusch V, Cedar H, Marcus M (1985) DNase I sensitivity in facultative and constitutive heterochromatin. Chromosoma 93:38–42
Stierhof YD, Schwarz H (1988) Immunoelectronmicroscopy on cryosections section permeability to specific antibodies, protein A-gold complexes and ferritin conjugated IgGs. Institute of Physics Conference Series no. 93 3:519–520
Stierhof YD, Schwarz H, Frank H (1986) Transverse sectioning of plastic-embedded immunolabeled cryosections: morphology and permeability to protein A-colloidal gold complexes. J Ultrastruct Res 97:187–196
Thiry M (1991a) DNase I-sensitive sites within the nuclear architecture visualized by immunoelectron microscopy. DNA Cell Biol 10:169–180
Thiry M (1991b) In situ nick-translation at the electron microscope level: a tool for studying the location of DNase I-sensitive regions within the cell. J Cytochem Histochem 39:871–874
Thiry M (1992a) Ultrastructural detection of DNA within the nucleolus by sensitive molecular immunocytochemistry. Exp Cell Res 200:135–144
Thiry M (1992b) Highly sensitive immunodetection of DNA on sections with exogeneous terminal deoxynucleotidyl transferase and non-isotopic nucleotide analogues. J Histochem Cytochem 40:411–419
Thiry M (1992c) New data concerning the functional organization of the mammalian cell nucleolus: Detection of RNA and rRNA by in situ molecular immunocytochemistry. Nucleic Acids Res 20:6195–6200
Thiry M (1993a) Ultrastructural distribution of DNA and RNA within the nucleolus of human Sertoli cells as seen by molecular immunocytochemistry. J Cell Sci 105:33–39
Thiry M (1993b) Immunodetection of RNA on ultrathin sections incubated with polyadenylate nucleotidyl transferase. J Histochem Cytochem 41:657–665
Thiry M (1993c) Differential location of nucleic acids within interchromatin granule clusters. Eur J Cell Biol 62:259–269
Thiry M (1994) Cytochemical and immunocytochemical study of coiled bodies in different cultured cell lines. Chromosoma 103:268–276
Thiry M (1995a) Nucleic acid compartmentalization within the cell nucleus by in situ transferase-immunogold techniques. Microsc Res Techn 31:4–21
Thiry M (1995b) Ultrastructural detection of nucleic acids by immunocytology. In: Morel G (ed) Visualization of nucleic acids. CRC Press. Boca Raton, pp 111–135
Thiry M (1995c) Behaviour of interchromatin granules during the cell cycle. Eur J Cell Biol (in press)
Thiry M, Goessens G (1986) Ultrastructural study of the relationships between the various nucleolar components in Ehrlich tumour and HEp-2 cell nucleoli after acetylation. Exp Cell Res 164:232–242
Thiry M, Goessens G (1991) Distinguishing the sites of pre-rRNA synthesis and accumulation in Ehrlich tumor cell nucleoli. J Cell Sci 99:759–767
Thiry M, Goessens G (1992) Where, within the nucleolus, are the rRNA genes located? Exp Cell Res 200:1–4
Thiry M, Puvion-Dutilleul F (1995) Differential distribution of single-stranded DNA, double-stranded DNA, and RNA in adenovirus-induced intranuclear regions of HeLa cells. J Histochem Cytochem (in press)
Thiry M, Thiry-Blaise L (1989) In situ hybridization at the electron microscope level: an improved method for precise localization of ribosomal DNA and RNA. Eur J Cell Biol 50:235–243
Thiry M, Scheer U, Goessens G (1991a) Localization of nucleolar chromatin by immunocytochemistry and in situ hybridization at the electron microscopic level. Electron Microsc Rev 4:85–110
Thiry M, Schoonbroodt S, Goessens G (1991b) Cytochemical distinction of various nucleolar components in insect cells. Biol Cell 72:133–140
Thiry M, Ploton D, Ménager M, Goessens G (1993) Ultrastructural distribution of DNA within the nucleolus of various animal cell lines or tissues revealed by terminal deoxynucleotidyl transferase. Cell Tissue Res 271:33–45
Vandelaer M, Thiry M, Goessens G (1993) Ultrastructural distribution of DNA with the ring-shaped nucleolus of human resting T lymphocytes. Exp Cell Res 205:430–432
Visa N, Puvion-Dutilleul F, Bacchellerie JP, Puvion E (1993) Intranuclear distribution of U1 and U2 SnRNAs visualized by high resolution in situ hybridization: revelation of a novel compartment containing U1 but not U2 SnRNA in HeLa cells. Eur J Cell Biol 60:308–321
Wassef M, Burglen J, Bernhard W (1979) A new method for visualization of preribosomal granules in the nucleolus after acetylation. Biol Cell 34:153–158
Wasylyk B, Thevenin G, Oudet P, Chambon P (1979) Transcription of in vitro assembled chromatin byEscherichia coli RNA polymerase. J Mol Biol 128:411–440
Williams M (1977) Quantitative methods in biology. In: Glauert A (ed) Practical methods in electron microscopy, vol 6. Elsevier North-Holland, Amsterdam, pp 1–216
Wolffe A, Drew H (1989) Initiation of transcription on nucleosomal templates. Proc Natl Acad Sci USA 86:9817–9821
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Thiry, M. New approaches to in situ detection of nucleic acids. Histochem Cell Biol 104, 81–95 (1995). https://doi.org/10.1007/BF01451570
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DOI: https://doi.org/10.1007/BF01451570