Oncology Probes

Preimplantation Genetic ScreeningFISH is the current gold standard to determine the chromosomal constitution of an embryo. In contrast tokaryotyping it can be used on interphase chromosomes, so that it can be applied on polar bodies, blastomeres andother single cell samples. FISH is therefore accepted as a routine method in preimplantation genetic screening(PGS) in determining chromosome aneuploidies prior to implanting an embyro and increases the success rate ofan IVF-mediated pregnancy.Preimplantation Genetic ScreeningMultiStar 24 FISHMultiStar 24 FISH consists of four DNA probe mixes eachhybridizing to six different chromosomes using six differentfluorochromes. It can be applied in lymphocytes, sperm andblastomeres. The fully optimized protocol consists of three15-30 minutes hybridizations followed by an overnight one. Inbetween the individual hybridizations, the preceding probesare washed off after imaging the results. The morphologyof the cell types is retained despite repeated denaturation,hybridization and post hybridization washes. Moreover theentire protocol can be completed within 24 hours, which fitsthe window for clinical PGS application. This novel methodeliminates the bottleneck perceived when using FISH byomitting key chromosomes relevant to implantation failurenot covered in a limited panel.Setup of the different layersAll DNA probes are labeled with PlatinumBright based on theUniversal Linkage System (ULS), KREATECH´s proprietary nonenzymaticlabeling technology capable of linking fluorescentlabels or haptens to any nucleic acid of interest.ColorMetaphase preparation from male lymphocyte cellsvisualizing all 24 chromosomes.Image kindly provided by Prof. D. Griffin, University of Kent, United KingdomThe probes required to provide information for 24 chromosomescomprise four panels each containing 6 chromosome-specificidentifier sequences each labelled with a different fluorochrome.The first 3 panels use centromeric sequences (Panel 1: chromosomes1,3,4,6,7,8; Panel 2: chromosomes 9,10,11,12,17,20 and Panel3: chromosomes 2,15,16,18,X,Y) . In contrast, Panel 4 used forthe final round of hybridization uses unique sequence probes forchromosomes 5,13,14,19,21,22 since centromeric sequenceswere not available for these chromosomes. A combination ofseparate probe denaturation for 10 minutes followed by a shortco-denaturation between probe and sample is used. Panels 1-3require short hybridization times (15 minutes) whereas Panel 4requires overnight hybridization.Label PlatinumBright 405 PlatinumBright 415 PlatinumBright 495 PlatinumBright 547 PlatinumBright 590 PlatinumBright 647MultiStar Panel 1 7 1 6 8 3 4MultiStar Panel 2 11 9 20 12 10 17MultiStar Panel 3 18 Y X 16 2 1566MultiStar Panel 4 19 5 21 22 13 14