Breast Surgery
Tjun Tang, Elizabeth O'Riordan, Stewart Walsh in Cracking the Intercollegiate General Surgery FRCS Viva, 2020
How is HER2 measured?Immunohistochemistry – antibodies directed against the HER2 protein are visualised with chromogenic detection. This is a widely available method. Score is 0, 1+, 2+ or 3+. There is a 10% false-negative rate. Equivocal results (2+) are retested by FISH.FISH is a quantitative measurement of gene amplification. It requires specialist equipment and is relatively expensive.CISH (chromogenic in situ hybridisation) and SISH (silver in situ hybridisation) are quantitative measurements using immunohistochemical techniques to visualise HER2 expression directly; CISH is lower cost than FISH, with similar sensitivity and specificity to the FISH technique.38
The N-Myc Oncogene in Pediatric Tumors: Diagnostic, Prognostic and Biological Aspects
John T. Kemshead in Pediatric Tumors: Immunological and Molecular Markers, 2020
As mentioned above, gene amplification is one of the mechanisms by which proto-oncogenes can become activated. In lower organisms, such as amphibians and Drosophila, selective increase in gene copy number (i.e., amplification) occurs normally during certain stages of development.17 Even in mammals, the phenomenon is not exclusively associated with the neoplastic process, but may be seen when cells are placed under a variety of selective conditions.12,18 The best known example of this is the amplification of the dihy-drofolate reductase gene, leading to drug-resistance in response to methotrexate exposure.19 At the cytological level, amplified genes are associated with characteristic karyotype abnormalities known as homogenously staining chromosomal regions (HSRs) and extrachro-mosomal double minute chromatin bodies (DMs). HSRs (so called because they tend to stain uniformly with Giemsa in G-band preparations) are relatively stable structures but DMs lack centromeres, are thus comparatively unstable and may disappear from cells altogether or participate in the formation of HSRs by chromosomal reintegration.18
Neoplasia
C. Simon Herrington in Muir's Textbook of Pathology, 2020
Gene amplification is a process whereby multiple copies of an oncogene are formed by reduplication. Transcription of these extra copies of the gene results in increased production of the oncoprotein, e.g. the N-MYC gene on chromosome 2 is greatly amplified in many cases of neuroblastoma, a rare childhood tumour of primitive neurons. The extra copies may be located on the correct chromosome, where they can be recognized as a ‘homogeneously staining region’, or as numerous extra chromosomal structures known as ‘double minutes’. The consequence in either case is of increased synthesis of the MYC protein, a transcription factor for genes involved in cell proliferation.
Loop-mediated isothermal amplification for Candida species surveillance in under-resourced setting: a review of evidence
Published in Expert Review of Molecular Diagnostics, 2022
Oloche Owoicho, Charles Ochieng’ Olwal, Edward Jenner Tettevi, Bernard Ortwer Atu, Ernest Uzodimma Durugbo
Typically, gene amplification occurs by repetition of two types of elongation reactions that occur via loop regions. Each of the inner primers has a sequence complementary to one chain of the amplification region at the 3’ terminal and is identical to the inner region of the same chain at the 5’ terminal. The elongation is sequentially repeated by DNA polymerase-mediated strand-displacement synthesis using the stem loop regions as a stage [17]. This method is premised on the principle of the production of lots of DNA amplification products with a mutually complementary sequence and an alternating repeated structure. LAMP amplifies a few copies of DNA to 109 copies within an hour under isothermal condition with great specificity (reviewed in [33]). The LAMP output is then visualized by the naked eye through magnesium pyrophosphate, a turbid by-product of the amplification. Amplification can also be visualized by adding different dyes like SYBR green, hydroxynaphthol blue, and calcein [27]. Moreover, other methods of detecting LAMP amplicons include the use of fluorogenic primers and probes, and heterogeneous or homogeneous particles, which have been well reviewed [20,21].
Inverse relationship between cagG-positive Helicobacter pylori status and risk of gastric ulcer
Published in British Journal of Biomedical Science, 2019
SZ Bakhti, N Raei, S Latifi-Navid, S Zahri, A Yazdanbod
Two hundred H. pylori isolates were obtained from gastric biopsy cultures of 145 (72.5%) patients with endoscopically defined non-atrophic gastritis, 27 (13.5%) with gastric ulceration and 28 (14.0%) with duodenal ulceration. Informed written consent was obtained from each participant, and the study was approved by the local research ethics committee. Biopsies were cultured and identified on selective Brucella agar plates supplemented with antibiotics, under microaerobic conditions at 37 °C for a maximum of 5–7 days. Colonies were identified as H. pylori according to standard criteria including negative Gram staining, typical cell morphology, and positive reactions to catalase, oxidase, and urease, as well as PCR amplification of H. pylori, 16S rDNA. Single colonies were isolated to ensure that each strain consisted of only a single genotype. DNA was extracted from H. pylori isolates with the Genomic DNA Purification kit (Fermentas, UK). The presence of the cagPAI genes was determined by PCR amplification and sequencing. The specific primers of each gene and their optimized annealing temperature, which were used for PCR amplification and sequencing, are listed in Table 1. The total volume of the reaction was 30 μL and included PCR buffer 3 μL, MgCl2 1 μL, dNTP mix 0.5 μL, primers (reverse and forward mixture) 1 μL, template DNA (5 μL) 25 ng, Taq DNA polymerase 0.2 μL, and distilled water 19.3 μL. The gene amplification conditions were previously described [2].
Dietary Pattern, Genomic Stability and Relative Cancer Risk in Asian Food Landscape
Published in Nutrition and Cancer, 2022
Razinah Sharif, Suzana Shahar, Nor Fadilah Rajab, Michael Fenech
Telomeres consist of a conserved hexanucleotide repeat sequence (TTAGGG) that caps the ends of chromosomes and protects them from recombining with each other and thus preventing chromosomal end-to-end fusions. Degradation of telomeres has been shown to lead to chromosomal instability, via telomere end fusions resulting in generation of abnormal dicentric chromosomes and breakage-fusion-bridge cycles within these abnormal chromosomes (75, 76). This leads to gene amplification and gene dosage imbalance which is an important risk factor for cancer. Accelerated telomere shortening and telomere end fusions can result in a persistent DNA damage response leading to cell cycle arrest and apoptosis. Although telomere shortening has been proposed as one of the fundamental mechanisms that determine chromosomal instability, and increased cancer risk, studies on the relationship between dietary factors and telomere biology have mainly focused on western diets such as the Mediterranean diet which is protective but knowledge remains limited with respect to Asian dietary patterns (77, 78).
Related Knowledge Centers
- DNA
- DNA Repair
- DNA Replication
- Molecular Evolution
- Retrotransposon
- Slipped Strand Mispairing
- Polyploidy
- Aneuploidy
- Gene
- Ectopic Recombination