For the present study, fluorescence in situ hybridisation (FISH) was employed for detection of TOP2A and chromosome 17 according to the manufacturer`s guidelines. Pretreatment was done using Histology FISH Accessory Kit, code K5799 (Dako). The probe mix (VYSIS TOP2A/CEP 17 FISH Probe Kit, code 03N89-020 Abbott Molecular Inc) was applied and denatured at 73°C for 5 min before hybridisation at 37°C overnight. For HER2 and chromosome 17, the HER2 CISH pharmDx Kit, code 109 (Dako), was used and immunostaining for ER (ER SP1 Cell Marqque 33 mg/mL 1:100) and PR (PR 16 Novocastra 360 mg/mL 1:400) was done in a DakoCytomation Autostainer Plus (Dako) using Dako REAL EnVision Detection System with Peroxidase/DAB+, Rabbit/Mouse, code K5007, as previously described.5 (link)
Protocol detail
Fluorescence In Situ Hybridization for Breast Cancer Markers
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Cell
Chromosome 17
Cish
Fluorescence in situ hybridisation
Her2
Hybridisation
Mouse
Peroxidase
Rabbit
Top2a
Corresponding Organization : St Olav's University Hospital
Other organizations : Norwegian University of Science and Technology
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Variable analysis
independent variables
- Probe mix (VYSIS TOP2A/CEP 17 FISH Probe Kit, code 03N89-020 Abbott Molecular Inc)
- HER2 CISH pharmDx Kit, code 109 (Dako)
- Immunostaining for ER (ER SP1 Cell Marqque 33 mg/mL 1:100) and PR (PR 16 Novocastra 360 mg/mL 1:400)
- Detection of TOP2A and chromosome 17
- Detection of HER2 and chromosome 17
- ER and PR immunostaining
- Histology FISH Accessory Kit, code K5799 (Dako)
- Pretreatment using Histology FISH Accessory Kit
- Probe mix denaturation at 73°C for 5 min
- Hybridisation at 37°C overnight
- Dako REAL EnVision Detection System with Peroxidase/DAB+, Rabbit/Mouse, code K5007
- Positive control: Not explicitly mentioned
- Negative control: Not explicitly mentioned
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