Nucleic Acids Research Advance Access published online on January 23, 2007
Nucleic Acids Research, doi:10.1093/nar/gkl1097
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Methods online polymorphism/ mutation detection and genomics |
Dry-reagent disposable dipstick test for visual screening of seven leukemia-related chromosomal translocations
1Department of Chemistry, University of Patras, Patras 26500, Greece, 2Hematology Division, Department of Internal Medicine, University of Patras Medical School, Patras, Greece 26500, 3Foundation for Research and Technology Hellas, Institute of Chemical Engineering and High Temperature Chemical Processes, (FORTH/ICE-HT), P.O. Box 1414, Patras 26504, Greece and 4Department of Chemistry, University of Athens, Athens 15771, Greece
*To whom correspondence should be addressed. Tel: +30 2610 996022; Fax: +30 2610 997118; E-mail: tkc{at}chemistry.upatras.gr
Received March 15, 2006. Revised December 1, 2006. Accepted December 1, 2006.
We report the first dry-reagent, disposable, dipstick test for molecular screening of seven chromosomal translocations associated with acute and chronic leukemia. The dipstick assay offers about 10 times higher detectability than agarose gel electrophoresis and, contrary to electrophoresis, allows confirmation of the sequence of the polymerase chain reaction (PCR) product by hybridization within a few minutes without the need of instrumentation. Biotinylated amplified DNA is hybridized with a dA-tailed probe and applied to the strip, which contains oligo(dT)-conjugated gold nanoparticles in dry form. Upon immersion of the strip in the appropriate buffer, the solution migrates and the hybrids are captured by immobilized streptavidin at the test zone generating a characteristic red line. The excess nanoparticles are captured by oligo(dA) strands immobilized at the control zone of the strip producing a second red line. We studied the: t(9;22)(q34;q11), t(15;17)(q22;q21), t(11;17)(q23;q21), t(5;17)(q32;q21), t(11;17)(q13;q21), t(8,21)(q22;q22) and inv(16)(p13;q22) that generate the BCR-ABL, PML-RARa, PLZF-RARa, NPM-RARa, NuMA-RARa, AML1-ETO and CBFß-MYH11 fusion genes, respectively. A single K562 cell was detectable amidst 106 normal leukocytes. A dipstick test was developed for actin, as a reference gene. The dipstick assay with appropriate probes can be used for identification of the fusion transcripts involved in the translocation.