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.