BCL-ABL M-bcr FusionQuant® Kits From Ipsogen

The M-bcr FusionQuant® Kit is used for the quantitative detection of M-bcr fusion transcripts on real-time thermal cyclers. Ipsogen has designed a very sensitive, reliable and easy to use kit. This real-time PCR kit is intended for the accurate quantification of BCR-ABL p210 transcripts in bone marrow or peripheral blood samples of ALL (Acute Lymphoid Leukemia) or CML (Chronic Myeloid Leukemia) patients previously diagnosed with a M-bcr fusion gene event. The results we obtain are used in monitoring the efficiency of treatment in patients undergoing therapy, and for minimum residual disease (MRD) follow-up to monitor disease relapse.

The protocol supplied with the kit is 22 pages and thoroughly describes all aspects of the assay, including intended use, background, technological principal, technological specifications, required reagents and instrumentation, instructions for use with different real-time PCR platforms, interpretation of results, troubleshooting guide, performance characteristics (non clinical/clinical studies) and references. The kit is standardized for use on all the major real-time instruments (including Applied Biosystems cyclers, Cepheid’s Smart Cycler, Corbett’s Rotor Gene, and Roche’s LightCycler). The kit contains the following components:
Three standards for House Keeping gene (ABL) – 10x3, 10x4 and 10x5 (50 ul each)
Five standards for fusion gene (M-bcr) – 10x1, 10x2, 10x3, 10x5 and 10x6 (50 ul)
Primer Probe mix for ABL (90 ul)
Primer probe mix for fusion gene (110 ul)

The kit has to be stored at -15°C to -25°C in a constant temperature freezer. Vortexing and centrifugation of the primer probe mix prior to use is very crucial; otherwise, it can lead to very high initial background. The kit comes with a long expiry date (~ 12 – 14 months from the date of purchase).

Total RNA (tRNA) extraction, conversion of tRNA into cDNA and cDNA detection on real-time PCR are all involved in reporting of a M-bcr sample. Reagents for tRNA extraction, cDNA conversion and real-time amplification have to be purchased separately. We use Qiagen kit (QIAamp RNA Blood Mini Kit) for extracting tRNA from whole blood and Takara’s cDNA PrimeScript First Strand cDNA Synthesis Kit for converting tRNA to cDNA. The real-time PCR instrument we use for the amplification and detection is the Cepheid SmartCycler. To set up the assay, we just need to put the PCR master mix (12.5 µl), primer probe mix (1 µl), the material to be quantified (sample cDNA/standard/water control) (5 µl), and finally, 6.5 µl of nuclease-free water to adjust the volume to 25 µl. Total run time is near 1hr 43 minutes on a SmartCycler.

In summary, the M-bcr FusionQuant® Kit offers an unparalleled fusion transcript detection through real-time PCR technology. This can be strongly recommended to anyone interested in a precise determination, quantification, and follow-up of 9:22 fusion chromosome.