Abstract
This paper describes a micro fluorescence in situ hybridization (μFISH)-based rapid detection of cytogenetic biomarkers on formalin-fixed paraffin embedded (FFPE) tissue sections. We demonstrated this method in the context of detecting human epidermal growth factor 2 (HER2) in breast tissue sections. This method uses a non-contact microfluidic scanning probe (MFP), which localizes FISH probes at the micrometer length-scale to selected cells of the tissue section. The scanning ability of the MFP allows for a versatile implementation of FISH on tissue sections. We demonstrated the use of oligonucleotide FISH probes in ethylene carbonate-based buffer enabling rapid hybridization within <1 min for chromosome enumeration and 10–15 min for assessment of the HER2 status in FFPE sections. We further demonstrated recycling of FISH probes for multiple sequential tests using a defined volume of probes by forming hierarchical hydrodynamic flow confinements. This microscale method is compatible with the standard FISH protocols and with the Instant Quality FISH assay and reduces the FISH probe consumption ∼100-fold and the hybridization time 4-fold, resulting in an assay turnaround time of <3 h. We believe that rapid μFISH has the potential of being used in pathology workflows as a standalone method or in combination with other molecular methods for diagnostic and prognostic analysis of FFPE sections.
Highlights
The most common cancer in females is breast cancer with 464 000 new cases alone in 2012, of which 131 000 cases were lethal.1 While the breast cancer incidence rate has been increasing, the death rate has decreased by 40% since the 1990s.2 This is likely due to early detection of the solid tumor linked to efforts in public health to improve awareness among women and routine screening
The mobility of fluorescence in situ hybridization (FISH) probes in the cell decreases with the increasing length, and it has been found that an optimal length for oligonucleotide FISH probes is 18–50 nucleotides
We introduced methods and protocols for rapid detection of an important breast cancer biomarker, human epidermal growth factor receptor 2 (HER2), in tissue sections
Summary
The most common cancer in females is breast cancer with 464 000 new cases alone in 2012, of which 131 000 cases were lethal. While the breast cancer incidence rate has been increasing, the death rate has decreased by 40% since the 1990s.2 This is likely due to early detection of the solid tumor linked to efforts in public health to improve awareness among women (at risk) and routine screening. The most common cancer in females is breast cancer with 464 000 new cases alone in 2012, of which 131 000 cases were lethal.. While the breast cancer incidence rate has been increasing, the death rate has decreased by 40% since the 1990s.2. This is likely due to early detection of the solid tumor linked to efforts in public health to improve awareness among women (at risk) and routine screening. The detection of breast cancer related biomarkers through genomic and transcriptomic analysis allows reclassification of the tumors into molecular subtypes such as Luminal A [ERþ, PRþ, and human epidermal growth factor 2 (HER2-)], Luminal B (low p53 mutation, ERþ, PRþ, and HER2þþ), Basal-like (Triple-negative), or HER2þþ [Fig. 1(a)].
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