Overview

Detection of genetic change for t(15;17) is important for the diagnosis of atypical variants of acute promyelocytic leukemia (APL) and for predicting the response to ATRA treatment.[1-4] For this purpose, morphological diagnosis in conjunction with cytogenetics and reverse transcrip-tion-polymerase chain reaction (RT-PCR) for promyelo-cytic leukemia (PML)/retinoic acid receptor alpha (RARA) is generally used. At follow-up after treatment of APL, it is not easy to decide upon remission by morphology. With respect to the discrimination between leukemic promyelocytes and regenerating promyelocytes, many points of discrimination have been suggested;[5] however, the situation is sometimes perplexing. A positive result for RT-PCR in patients with PML after chemotherapy does not mean a persistence of malignant cells, because differentiated leukemic cells by ATRA treatment still harbor the PML/RARA gene rearrangement. Actually, although the majority of APL patients achieve morphological remission after complete remission induced by ATRA, PML/RARA transcripts remain detectable in all cases.[6-8] Hence serial quantitation of leukemic cells harboring the PML/RARA rearrangement is essential in the assessment of complete remission after consolidation.1-9-1

For the accurate and quantitative detection of minimal residual disease (MRD), real-time RT- PCR is available but is not yet used widely. Furthermore, persistence of PCR positivity in long-term remission without recurrence has been reported,[10] which means those clones with PCR+ do not always have leukemogenic potential. Therefore the detection of PML/RARA fusion by FISH in the initial follow-up after treatment is very helpful. In addition, there are many factors that can influence the results of PCR, such as the quality of the mRNA, the timing of sampling, and the sensitivity of the RT-PCR assay. Although the sensitivity of FISH (10" 2-10"3) is lower than that of PCR (10" 5-10"6), the results of FISH are more precise, highly concordant between laboratories, and not influenced by other factors.[11] The only factor that can influence the interpretation of results is the different cutoff value used by laboratories. The scoring of fusion and split signals in FISH may vary by individual, resulting in different cutoff values among laboratories.

Single FISH probes for the PML/RARA fusion gene are commercially available, and the scheme of the PML/ RARA probe involves the use of two probes: one from each of the fusion genes, differentially labeled. However, chance of colocalization of differently colored signals in normal specimens can produce some levels of false-positive nuclei with some traditional, two-color, come-together FISH translocation probes. The high normal range of probes with design of the PML/RARA probes is inherent in that design, but extra signal or dual-fusion PML/RARA probes are not available at the present time. This false-positive fusion encountered in normal cells makes it difficult to detect minimal residual disease (MRD) quantitatively, especially during follow-up. If we use RARA probes that are located in the proximal and distal sides of the breakpoint of the RARA gene, the signals would be split, resulting in one green signal, one orange signal, and one fusion signal (1G1O1F), and these probes would enable us to detect the translocations of the RARA gene with any partner chromosomes. The LSI RARA Rearrangement Probe (Vysis, Downers Grove, IL, USA) was originally designed to detect nuclei that have variant rearrangements involving the RARA gene and genes other than PML. However, this probe may also provide better information in the detection of nuclei that have t(15;17), because of its break-apart design.

We performed PML/RARA FISH and RARA break-apart FISH on bone marrow specimen of patients to investigate whether the RARA break-apart probe can overcome the disadvantage of the false-positive fusion signal of the single-FISH (S-FISH) for PML/RARA, and to evaluate the efficiency of RARA break-apart FISH in the diagnosis and monitoring of APL.

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Getting Started With Dumbbells

Getting Started With Dumbbells

The use of dumbbells gives you a much more comprehensive strengthening effect because the workout engages your stabilizer muscles, in addition to the muscle you may be pin-pointing. Without all of the belts and artificial stabilizers of a machine, you also engage your core muscles, which are your body's natural stabilizers.

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