Types of FISH Probes

Types of FISH Probes

• Fluorescent in situ hybridization (FISH) uses fluorescently labeled DNA probes to pinpoint specific DNA sequences within chromosomes or cells, identifying genetic alterations.
• Different probes target different chromosomal regions.
• Specific probes bind to unique DNA sequences for a particular target region.
• Centromeric probes target the centromere region of chromosomes.
• Telomeric probes target the telomere regions of chromosomes.
• Whole chromosome paints use whole chromosomes as target sequences.
• Locus-specific probes target specific DNA sequences within genes.
• Dual-color probes label two different DNA sequences with differing fluorophores, visualizing the relative positions of targeted genes/regions.
• Multicolor probes label multiple DNA sequences with various fluorophores for comprehensive genomic analysis, such as visualizing all chromosomes simultaneously.

Probe Types and Applications

• Spectral karyotyping (SKY) probes paint all chromosomes, providing a detailed view of chromosome structure and arrangement.
• Comparative genomic hybridization (CGH) compares a sample's DNA with a reference to identify chromosomal alterations like deletions or duplications.
• Array comparative genomic hybridization (aCGH) analyzes genome-wide chromosomal aberrations with high resolution, detecting smaller changes than CGH.
• Break-apart probes contain two fluorophore-labeled DNA fragments, hybridizing to adjacent but separate regions of a gene. A break in the gene prevents probe binding, crucial for detecting physical gene breaks.
• Dual-color break-apart probes detect gene breaks and translocations by identifying the specific gene and its breakpoints.
• Dual fusion probes identify gene relocation, fusion, or rearrangements on a chromosome.
• Bacterial artificial chromosome (BAC) probes analyze larger chromosomal regions, suitable for specific location analysis.

Probe Design Considerations

• Probe specificity is vital; the probe sequence must uniquely target the desired DNA region to minimize nonspecific binding.
• Longer probes are more robust and stable during hybridization.
• Fluorescent labels like FITC, ROX, Cy3, and Cy5 are used for probe visualization.
• Proper probe concentration ensures optimal hybridization.
• Probe purity prevents false-positive signals due to contaminants.
• Probe synthesis methods affect specificity.

Differences between FISH Probe Types

• Different probes target different chromosomal regions (centromeres, telomeres, specific genes).
• Different probes detect different genomic alterations (disruptions, fusions, gross rearrangements).
• Resolution varies; whole chromosome paints have lower resolution than locus-specific probes or break-apart probes.
• Probes use various fluorescent dyes for labelling, enabling color-coded identification of specific chromosomal regions in techniques like spectral karyotyping.
• Length and specificity influence efficient target identification.
• Probes target different sizes of genomic regions, from specific genes to whole chromosomes.

Practical Applications of FISH Probe Types

• Clinical diagnosis utilizes FISH to detect chromosomal abnormalities in cancer cells, genetic disorders, prenatal diagnosis, and to pinpoint specific translocations.
• Research investigates chromosome structure, gene function, and genome organization via FISH.
• Genetic screening employs FISH for early detection of genetic abnormalities in embryos and fetuses.
• Cancer research uses FISH to diagnose and monitor cancer progression and treatment response, understanding cancer development, progression, and resistance mechanisms.