Fluorescence In Situ Hybridization
In situ hybridization uses complementary base sequences between single strands of nucleic acid molecules to complement and pair radioactive or non radioactive foreign nucleic acids (i.e. probes) with DNA or RNA to be tested on tissues, cells or chromosomes to synthesize nucleic acid hybridization molecules. The nucleic acid to be tested is detected in tissues A position on a cell or chromosome. Fluorescence in situ hybridization (FISH) is a non radioactive molecular cellular genetic technology developed on the basis of radioactive in situ hybridization. The in situ hybridization method using fluorescent markers instead of isotopic markers is a molecular diagnostic technology using Fluorescent-labeled DNA probes to detect specific DNA sequences in cells, tissues and tumors.
Figure 1. Fluorescent in situ hybridization (Hao, et al. 2013).
Unlike other techniques used to study chromosomes, FISH is flexible and does not need to be detected in dividing cells. Therefore, it can be used to detect whether genes or chromosomes are abnormal. It is usually used for genetic counseling, medical and variety identification, and DNA specific function research. Relying on advanced experimental platform and professional research team, CD Biosciences can provide comprehensive and customized fluorescence in situ hybridization services to help your scientific research work. If you have any needs, please feel free to contact us.
Principle of Fluorescence In Situ Hybridization
The basic components of FISH are a DNA probe and a target DNA sequence located by it. Before hybridization, DNA probes are labeled by a variety of techniques, such as random primer labeling, nick translation, and PCR amplification. The following two labeling methods are commonly used: direct labeling and indirect labeling (Fig. 2). For the direct labeling method, nucleotides that have been directly altered to contain a fluorophore are used; however, in indirect labeling, the probe is labeled with altered nucleotides with haptens. Thereafter, the labeled probe and the target DNA are denatured, and the denatured probe is paired with the target DNA during the annealing of the cDNA sequence. If the probe is indirectly labeled, an additional step is required to observe the non-fluorescent hapten using immunological or enzymatic detection systems. Therefore, directly labeled probes are faster, and indirect labeling provides sufficient signal amplification by using multiple antibody membranes.
Figure 2. Principle of Fluorescence in situ hybridization (Mohanty, et al. 2020).
Different Types of FISH and Their Functions
|ACM-FISH||In sperm cell, structural and numerical chromosomal abnormalities can be detected|
|CARD-FISH||Amplification of signal which is obtained by peroxidase activity|
|CAT-FISH||Expression of genes patterns in brain|
|CO-FISH||The orientation of tandem repeats in the centromeric regions of chromosomes|
|CB-FISH||The cytological analysis of micronucleation and aneuploidy|
|COD-FISH||Quantification of gene copy number and the protein amount|
|D-FISH||Detection of BCR/ABL fusion in chronic myeloid leukemia (CML)|
|DBD-FISH||Any sites of DNA damage/breakage in the sample genome|
|Fiber-FISH||Mapping of genes and chromosomal regions on fibers of chromatin or DNA|
|Flow-FISH||Visualize and measure the length of telomere|
|Harlequin-FISH||Cell cycle-controlled chromosome analysis in human lymphocytes|
|Immuno-FISH||Both DNA and proteins can be analyzed in the same sample|
|M-FISH||Facilitating the analysis of complex chromosomal rearrangement|
|ML-FISH||Identifying multiple microdeletion syndromes in patients|
|PCC-FISH||Chromosome damage after irradiation|
|Q-FISH||Determining the repeated number of telomere on a specific chromosome|
|QD-FISH||Human metaphase chromosomes, human sperm cells, bacterial cells, and also to detect subcellular mRNA distribution in tissue sections|
|Raman-FISH||For characterizing of chromosomes and chromosome amplifications in cancer|
|RING-FISH||Identification of individual genes and detection of halo appearance from fluorescence signals at the bacterial cell at periphery|
|RNA-FISH||Allelic-specific expression in per cell basis|
|T-FISH||Mapping of gene loci and looking for specific transcripts in cells|
CARD-FISH: catalyzed reporter deposition FISH; CAT-FISH: capture antibody targeted detection FISH; CB-FISH: cytochalasin B FISH; CO-FISH: cyotochrome orientation FISH; COD-FISH: chromosome orientation and direction FISH; D-FISH: dual color FISH; DBD-FISH: deoxyribonucleic acid breakage detection FISH; DNA: deoxyribonucleic acid; M-FISH: multiple spectral karyotyping FISH; ML-FISH: multilocus FISH; mRNA: messenger ribonucleic acid; PCC-FISH: premature chromosome condensation FISH; Q-FISH: quantitative FISH; QD-FISH: quantum dots FISH; RNA: ribonucleic acid; T-FISH: tissue FISH
Applications of Fluorescence In Situ Hybridization
- Detection of numerical and chromosomal abnormalities
- Monitoring the effects of therapy
- Detection of early relapse or minimal residual disease
- Identification of the lineages of neoplastic cells
- Examination of the karyotypic pattern of interphase cells, including nondividing or terminally differentiated cells
- Detection of gene deletions and gene amplification
- Hao, Ming, et al. "Production of hexaploid triticale by a synthetic hexaploid wheat-rye hybrid method." Euphytica 193.3 (2013): 347-357.
- Mohanty, Swati Sucharita, Chita Ranjan Sahoo, and Rabindra Nath Padhy. "Role of hormone receptors and HER2 as prospective molecular markers for breast cancer: An update." Genes & Diseases (2020).
- Gozzetti, Alessandro, and Michelle M. Le Beau. "Fluorescence in situ hybridization: uses and limitations." Seminars in hematology. Vol. 37. No. 4. WB Saunders, 2000.
- Ratan, Zubair Ahmed, et al. "Application of fluorescence in situ hybridization (FISH) technique for the detection of genetic aberration in medical science." Cureus 9.6 (2017).
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