Chapter 14
PROTOCOL
Nucleus Extraction from Cryofixed Tissue
BERND RAUTENSTRAUSS and THOMAS LIEHR
Introduction
The advent of fluorescence in situ hybridization (FISH) rendered it possible to obtain "cytogenetic information" from interphase nuclei (Langer
et aI., 1981, Pinkel et al, 1986). Karyotypic analysis by direct demonstration
of DNA sequences in interphase nuclei (interphase cytogenetics or interphase FISH) can be applied to a wide variety of cellular material, including
cytogenetic preparations, fresh material (touch-preparations), cryofIxed
and paraffIn-embedded tissue. In contrary to metaphase FISH, only cosmids, PI-clones, BACs, YACs and satellite probes are suitable for interphase FISH. Detection and quantitation of structural chromosome aberrations, of microdeletions or microduplications and oncogene amplifIcation or deletion of tumor suppressor genes in solid tumors thus became
possible.
To date, fresh tumor material (e.g. Liehr et al., 1994) or formalin-fIxed
and subsequently paraffin-embedded and sectioned tissue (e.g. Neubauer
et aI., 1994) has mainly been used for interphase cytogenetic studies. Formalin-fIxed/paraffIn-embedded tissue is readily available, as this kind of
tissue fIxation is the most common standard technique in clinical practice.
However, there are some disadvantages in connection with this material
and an increasing number of laboratories now collect archival formalinfixed/paraffIn-embedded and cryofIxed tissue samples.
Formalin-fIxed/paraffIn-embedded tissue is, for example, not suitable
for all kinds of immunohistochemical approaches, as specifIc antigens
may be destroyed during the fIxation procedure. Moreover, if formalin
~ Bernd Rautenstrauss, Institut fUr Humangenetik, Schwabachanlage 10, Erlangen,
91054, Germany (phone +49-9131-8532352; fax +49-9131-209297;
e-mail berndwr@humgenet.uni-erlangen.de)
Thomas Liehr, Institut fUr Humangenetik, Kollegiengasse 10, lena, 07743, Germany
PROTOCOL
Nucleus Extraction from Cryofixed Tissue
BERND RAUTENSTRAUSS and THOMAS LIEHR
Introduction
The advent of fluorescence in situ hybridization (FISH) rendered it possible to obtain "cytogenetic information" from interphase nuclei (Langer
et aI., 1981, Pinkel et al, 1986). Karyotypic analysis by direct demonstration
of DNA sequences in interphase nuclei (interphase cytogenetics or interphase FISH) can be applied to a wide variety of cellular material, including
cytogenetic preparations, fresh material (touch-preparations), cryofIxed
and paraffIn-embedded tissue. In contrary to metaphase FISH, only cosmids, PI-clones, BACs, YACs and satellite probes are suitable for interphase FISH. Detection and quantitation of structural chromosome aberrations, of microdeletions or microduplications and oncogene amplifIcation or deletion of tumor suppressor genes in solid tumors thus became
possible.
To date, fresh tumor material (e.g. Liehr et al., 1994) or formalin-fIxed
and subsequently paraffin-embedded and sectioned tissue (e.g. Neubauer
et aI., 1994) has mainly been used for interphase cytogenetic studies. Formalin-fIxed/paraffIn-embedded tissue is readily available, as this kind of
tissue fIxation is the most common standard technique in clinical practice.
However, there are some disadvantages in connection with this material
and an increasing number of laboratories now collect archival formalinfixed/paraffIn-embedded and cryofIxed tissue samples.
Formalin-fIxed/paraffIn-embedded tissue is, for example, not suitable
for all kinds of immunohistochemical approaches, as specifIc antigens
may be destroyed during the fIxation procedure. Moreover, if formalin
~ Bernd Rautenstrauss, Institut fUr Humangenetik, Schwabachanlage 10, Erlangen,
91054, Germany (phone +49-9131-8532352; fax +49-9131-209297;
e-mail berndwr@humgenet.uni-erlangen.de)
Thomas Liehr, Institut fUr Humangenetik, Kollegiengasse 10, lena, 07743, Germany