Signature®LTx Leukemia Translocation Panel v2.0 (RUO)*
4 Diseases • 9 Translocations • 12 Fusion Transcripts • 1 Well
Use Signature® LTx v2.0 for the rapid detection and identification of
the fusion transcripts from common chromosome translocations/abnormalities associated
with CML, ALL, APL and AML*. Signature® LTx v2.0 utilizes both multiplex
RT-PCR and automated multiplex detection on the Luminex® 100™
IS or 200™ System.
Signature® LTx v2.0 provides comprehensive information in a single well
reducing the need for and cost of maintaining and training on numerous platforms
and assays.
Figure 1. Overview of the Signature LTx v2.0 assay principle and workflow
(click to enlarge).
Signature® LTx v2.0 has a simple workflow that requires minimal hands-on
time. The assay contains a single transfer step of amplified products directly to
the bead mix, without intervening purifications. The average time to results is
approximately 6 hours for up to 24 reactions per kit.
Table 2: Simultaneous Detection of 12 Leukemia-Associated Fusion Transcripts
and Endogenous Control GAPDH† (click to enlarge).
Signature® LTx Negative, Positive and No Target (NTC) controls were analyzed
together with 400 ng of total RNA specimens purified from 8 different translocation
positive leukemic cell lines. Qualitative results are measured by MFI (mean fluorescent
intensity). MFI values greater than or equal to 350 MFI signify positive detection
for a specific fusion transcript. A sample is considered negative if the MFI value
is below 350 MFI and the GAPDH signal is greater than or equal to 1,000 MFI. Specific
fusion transcripts were detected in specimens undiluted (100%) or diluted 1:100
(1%) into a background of 400 ng of translocation negative leukemic cell line RNA
(HL-60). This analytical sensitivity is equivalent to the detection of 1 fusion
transcript positive cell in a background of at least 100 fusion transcript negative
cells. No cross reactivity between the various targets was observed and MFI values
for all non-target probes were reproducibly below the cut-off (350 MFI).
Figure 2: Serial Dilution of Cell line total RNA† (click
to enlarge).
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Fusion transcripts were prepared by in vitro transcription (IVT) and spiked into
a background of total RNA purified from a translocation negative leukemic cell line.
In this experiment, 4 different IVT RNAs were serially diluted into a constant background
of 400 ng of HL-60 total RNA. Only MFI results for PML/RARα (L or S form), inv(16)
type D, MLL/AF4 (e10/e4) and the GAPDH probes are shown. Each fusion transcript
target was detected specifically in specimens containing 2x106, 2x105
or 2x104 copies (labeled 2M, 200K and 20K respectively in the figure).
The MFI obtained for the fusion transcripts and the endogenous control GAPDH transcript
were in the same range as those obtained with cell line RNA specimens.
Attributes for Signature® LTx* v2.0:
* Research Use Only. Not intended for use in diagnostic procedures.
† Preliminary research data, the performance characteristics
of this assay are not yet established.
References
Gabert J, Beillard E, van der Velden VH, Bi W, Grimwade D, Pallisgaard N, Barbany
G, Cazzaniga G, Cayuela JM, Cave H, Pane F, Aerts JL, De Micheli D, Thirion X, Pradel
V, Gonzalez M, Viehmann S, Malec M, Saglio G, van Dongen JJ. Standardization and
Quality Control Studies of “Real-Time” Quantitative reverse Transcriptase Polymerase
Chain Reaction of Fusion Gene Transcripts for Residual Disease Detection in Leukemia
– A Europe Against Cancer Program. Leukemia. 17(12):2318-57. 2003.
Salto-Tellez M, Shelat SG, Benoit B, Rennert H, Carroll M, Leonard DG, Nowell P,
Bagg A. Multiplex RT-PCR for the Detection of Leukemia-Associated Translocations:
Validation and Application to Routine Molecular Diagnostic Practice. J Mol Diagn.
5(4):231-6. 2003.
Bock O, Reising D, Kreipe H. Multipilex RT-PCR for the Detection of Common BCR-ABL
Fusion Transcripts in Paraffin-Embedded Tissues from Patients with Chronic Myeloid
Leukemia and Acute Lymphoblastic Leukemia. Diagn Mol Pathol. 12(3):119-23. 2003.
For more information, please contact orders@asuragen.com.