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Zusammenfassung

Die in situ-Hybridisierung (ISH) stellt eine Methode dar, die simultan den Nachweis spezifischer Genabschnitte der zellulären Desoxyribonucleinsäure (DNA) oder deren, auf Botenmessenger (m)-Ribonucleinsäure (RNA)-Niveau exprimierter Transkripte und die Beurteilung der Morphologie jener Zellen erlaubt, die über die entsprechenden Gensequenzen oder deren Transkripte verfügen. Aus dieser Definition resultieren die wesentlichen Vorteile und Indikationen dieser Technik (Tabelle 1), aber auch das Anforderungsprofil an die methodischen Details. Die üblichen Blotting-Verfahren von DNA (Southern) und RNA (Northern) liefern den Nachweis von spezifischen Genen oder ihrem Expressionsniveau an Gesamtextrakten von Geweben. Dies hat den Vorteil, daß die extrahierten, am Blot fixierten Nukleinsäuren extrem aggressiv behandelt werden, unspezifische Reaktionen der meist radioaktiv markierten Proben durch Abtrennung von Zellorganellen, ribosomaler (r)-RNA, Transfer (t)-RNA und Protein vermieden werden können. Im übrigen erlaubt die Bestimmung der jeweiligen Molekulargewichte der nachgewiesenen Reaktionspartner der Probe eine zuverlässige Bestimmung der Spezifität des Reaktionsprofils. Die Quantifizierung der Banden kann relativ einfach densitometrisch oder mittels Image-Analyse-Systemen erfolgen (s. u.). Folgende Nachteile dieser Methoden skizzieren zugleich Vorteile (Tabelle 1) und Einsatzspektrum (Tabelle 2) von in situ-Hybridisierungstechniken.

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© Springer-Verlag Berlin Heidelberg 1994

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  • R. Greil

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