Interphase in situ hybridization to disaggregated and intact tissue specimens of prostatic adenocarcinomas

Abstract

A comparative study was performed of interphase in situ hybridization (ISH) to deparaffinized 4-gm tissue sections and nuclear suspensions from eight prostatic adenocarcinomas, as well as one normal prostatic control. Whole nuclear suspensions were derived from the same tumor areas to evaluate differences of ISH to truncated versus whole nuclei. DNA probes specific for the centromeres of chromosome l, 7, 8, 10, and Y were used for detection of numerical chromosomal changes and aneuploidy. In six adenocarcinomas chromosome aberrations (+7, +8, -8, -10, -Y) were seen. However, ISH to sections revealed focal aberrations (-10, -Y) in four cases that could not be distinguished in the suspensions. Chromosomal alterations occurring in larger tumor areas were also detected in the nuclear suspensions. Chromosome copy number changes, especially gains, were better discriminated in the nuclear suspensions. The rate of ISH aneuploidy seen in nuclear suspensions corresponded with that observed in the tissue sections (P<0.01). Ploidy patterns as assessed by ISH to sections and nuclear suspensions were in concordance with DNA flow cytometry (both P<0.001). We conclude that both section and suspension ISH were able to accurately detect aneuploidy and numerical chromosomal aberrations occurring in larger histological areas. However, section ISH was also capable of revealing (small) focal cytogenetic abnormalities, due to a precise analysis of only target cells. Focal abnormalities were not detected by suspension ISH, probably due to an admixture of non-aberrant tumor cells and stromal elements.

PLAG1 gene alterations in salivary gland pleomorphic adenoma and carcinoma ex-pleomorphic adenoma: a combined study using chromosome banding, in situ hybridization and immunocytochemistry

“…CISH analysis was performed as previously described by Alers et al 25 CISH probes were labelled with fluorescein (Amersham Pharmacia), by random priming, as described above. Probe detection was performed using horseradish peroxidase antifluorescein (Roche) and H 2 O 2 -diaminobenzidine.…”

Longitudinal evaluation of cytogenetic aberrations in prostatic cancer: tumours that recur in time display an intermediate genetic status between non-persistent and metastatic tumours

“…A more detailed description of the probes used is given in refs 17, 25, and refs cited therein. Selection criteria were based on previous studies 16,17 and other literature data considering cytogenetic aberrations in prostatic tumours. 8,9,21,22 The (peri)centromeric repetitive satellite DNA probes were labelled with biotin-14-dATP by nick translation of complete plasmid DNA according to the manufacturer's directions (BRL, Gaithersburg, MD, U.S.A.).…”

“…A CGH study 15 performed on regional lymph node and bone metastases showed frequent gain of 8q, as well as loss of 8p, 10q, 13q, 16q and 17p sequences in over half of cases. Interphase in situ hybridization (ISH) to nuclear suspensions, touch preparations, and paraffin sections of both prostatic tumours and precursor lesions (PIN) revealed numerical aberrations of chromosomes 7, 8, 10, 16, 17, 18, X, and Y, [16][17][18][19] as well as loss of sequences in the 8p12-p22 region. 20 Furthermore, fluorescent ISH (FISH) studies of nuclear suspensions or touch imprints of mostly primary prostatic tumours suggested that alterations of chromosome 7 and/or 8 may be prognostic markers predicting unfavourable outcomes in prostate cancer.…”

Comparative FISH analysis of numerical chromosome 7 abnormalities in 5-μm and 15-μm paraffin-embedded tissue sections from prostatic carcinoma

“…From these sections, the corresponding tumor areas were selectively cut out with a fine scalpel under microscopic control. For disaggregation of tumor nuclei, the protocol of Alers et al (1995a) was modified. Briefly, after deparaffination and rehydration, samples were digested in a pronase E solution (0.05% in phosphatebuffered saline, PBS, pH 7.0) for 2 h at 37°C, with vigorous vortexing every 15 min.…”

“…Several studies have demonstrated the reliability of this technique for an evaluation of numerical chromosome aberrations (Alers et al 1995a;Bandyk et al 1994;Herrington et al 1995;Micale et al 1992). To enable a correlation of FISH analysis and histomorphological patterns, the method was modified for its application on routine paraffin-embedded tissue sections (van Dekken et al 1992;Dhingra et al 1994;Hopman et al 1991;Kim et al 1993).…”