Non-fluorescent chromosome painting using the peroxidase/diaminobenzidine (DAB) reaction.
OBJECTIVE
To develop and validate non-fluorescent chromosome painting for bright-field microscopy using the peroxidase/diaminobenzidine (DAB) reaction.


METHODS
Peripheral blood lymphocytes were taken from patients with uterine cancer who had received heavy-ion radiation therapy. Chromosome slides were treated with RNase and pepsin, denatured mildly, hybridized with a biotinylated DNA probe specific for whole-chromosome 4 and stained using the peroxidase/DAB reaction with an avidin-biotin amplification. The slides were analysed under a bright-field microscope and an atomic force microscope. The detection rate of chromosome aberrations by DAB painting was compared with that obtained by dual analysis of Giemsa staining and FISH painting.


RESULTS
When chromosomes 4 were painted, 11.5% of unstable aberrations were detected by DAB painting, while 10.8% of them were found by dual analysis of Giemsa staining and FISH painting.


CONCLUSIONS
A DAB painting method that can effectively detect rearranged aberrations was established. It has advantages over FISH painting: the preparations can be analysed by bright-field microscope, can be preserved permanently and are suitable for analysis by an automated system.
Kanda
R
R
Division of Radiobiology and Biodosimetry, National Institute of Radiological Sciences, Chiba, Japan.
Suzuki
M
M
Minamihisamatsu
M
M
Furukawa
A
A
Odaka
T
T
Hayata
I
I
D015100:3,3'-Diaminobenzidine; D000368:Aged; D002869:Chromosome Aberrations; D002894:Chromosomes, Human, Pair 4 / Q000528:radiation effects*; D015342:DNA Probes; D005260:Female; D006801:Humans; D017403:In Situ Hybridization; D008214:Lymphocytes / Q000528:radiation effects / Q000648:ultrastructure; D008853:Microscopy / Q000379:methods; D008875:Middle Aged; D010544:Peroxidases; D014594:Uterine Neoplasms / Q000097:blood / Q000235:genetics / Q000532:radiotherapy
D015342:DNA Probes; D015100:3,3'-Diaminobenzidine; D010544:Peroxidases