Abstract:
:We present the results of a novel method developed for evaluation of in situ amplification, a molecular genetic method at the cellular level. Reverse transcription polymerase chain reaction (RT-PCR) was used to study bcr-abl transcript levels in individual cells from patients with chronic myelogenous leukaemia (CML). After hybridizing a fluorochrome-labelled probe to the cell-bound RT-PCR product, bcr-abl mRNA-positive cells were determined using image analysis. A dilution series of bcr-abl-positive BV173 into normal cells showed a good correlation between expected and actual values. In 25 CML samples, the percentage of in situ PCR-positive cells showed an excellent correlation with cytogenetic results (r = 0.94, P < 0.0001), interphase fluorescence in situ hybridization (FISH) (r = 0.95, P = 0.001) and hypermetaphase FISH (r = 0.81, P < 0.001). The fluorescence intensity was higher in residual CML cells after interferon (IFN) treatment than in newly diagnosed patients (P = 0.004), and was highest in late-stage CML resistant to IFN therapy and lowest in CML blast crisis (P = 0.001). Mean fluorescence values correlated with bcr-abl protein levels, as determined by Western blot analysis (r = 0.62). Laser scanning cytometry allowing automated analysis of large numbers of cells confirmed the results. Thus, fluorescence in situ PCR provides a novel and quantitative approach for monitoring tumour load and bcr-abl transcript levels in CML.
journal_name
Br J Haematoljournal_title
British journal of haematologyauthors
Pachmann K,Zhao S,Schenk T,Kantarjian H,El-Naggar AK,Siciliano MJ,Guo JQ,Arlinghaus RB,Andreeff Mdoi
10.1111/j.1365-2141.2001.02510.xsubject
Has Abstractpub_date
2001-03-01 00:00:00pages
749-59issue
3eissn
0007-1048issn
1365-2141pii
bjh2610journal_volume
112pub_type
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journal_title:British journal of haematology
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