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Questions and Answers
What is primarily analyzed in fluorescence microscopy?
What is primarily analyzed in fluorescence microscopy?
Which type of microscope provides better resolution in fluorescence imaging by using optical sectioning?
Which type of microscope provides better resolution in fluorescence imaging by using optical sectioning?
What type of light typically excites the electrons in fluorescence microscopy?
What type of light typically excites the electrons in fluorescence microscopy?
Which of the following is NOT a feature of epifluorescence microscopy?
Which of the following is NOT a feature of epifluorescence microscopy?
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What does fluorescence microscopy use to generate an image?
What does fluorescence microscopy use to generate an image?
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What type of light sources are commonly used in fluorescence microscopy?
What type of light sources are commonly used in fluorescence microscopy?
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Which application of fluorescence is not commonly associated with cytogenetic analysis?
Which application of fluorescence is not commonly associated with cytogenetic analysis?
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What advantage does Fluorescence in situ Hybridization (FISH) have over routine chromosome analysis?
What advantage does Fluorescence in situ Hybridization (FISH) have over routine chromosome analysis?
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What imaging system is mentioned as part of cytogenetics work in the lab?
What imaging system is mentioned as part of cytogenetics work in the lab?
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Which feature is associated with imaging systems in fluorescence microscopy?
Which feature is associated with imaging systems in fluorescence microscopy?
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What is one of the primary uses of fluorescence in situ hybridization (FISH)?
What is one of the primary uses of fluorescence in situ hybridization (FISH)?
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Which of the following techniques involves multicolor fluorochrome banding?
Which of the following techniques involves multicolor fluorochrome banding?
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What impact does FISH have on the turnaround time of patient results?
What impact does FISH have on the turnaround time of patient results?
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What is the primary difference between DIRECT FISH and INDIRECT FISH techniques?
What is the primary difference between DIRECT FISH and INDIRECT FISH techniques?
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In the context of FISH detection, what does Tyramide signal amplification (T5A) enable?
In the context of FISH detection, what does Tyramide signal amplification (T5A) enable?
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Which of the following types of FISH probes is specifically designed to label repetitive sequences?
Which of the following types of FISH probes is specifically designed to label repetitive sequences?
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What is the purpose of denaturation in the FISH process?
What is the purpose of denaturation in the FISH process?
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What type of FISH probe is designed to target the centromere of chromosomes?
What type of FISH probe is designed to target the centromere of chromosomes?
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What visual pattern does dual color dual fusion fluorescent in situ hybridization (FISH) typically produce?
What visual pattern does dual color dual fusion fluorescent in situ hybridization (FISH) typically produce?
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How does the use of Horseradish peroxidase contribute to the Tyramide signal amplification process in mRNA detection?
How does the use of Horseradish peroxidase contribute to the Tyramide signal amplification process in mRNA detection?
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What does the term 'break apart' refer to in break apart FISH probes?
What does the term 'break apart' refer to in break apart FISH probes?
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What is the primary function of the BRCA1 protein?
What is the primary function of the BRCA1 protein?
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What percentage of breast cancer cases in the United States is associated with abnormal BRCA1 or BRCA2 genes?
What percentage of breast cancer cases in the United States is associated with abnormal BRCA1 or BRCA2 genes?
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Which test measures the presence of protein receptors or gene copy number for HER2 status?
Which test measures the presence of protein receptors or gene copy number for HER2 status?
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What does FISH specifically involve when analyzing genetic material?
What does FISH specifically involve when analyzing genetic material?
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What is the location of the BRCA1 gene on the chromosome?
What is the location of the BRCA1 gene on the chromosome?
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How does an abnormal BRCA1 or BRCA2 gene affect women's risk of breast cancer?
How does an abnormal BRCA1 or BRCA2 gene affect women's risk of breast cancer?
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What color indicates the visualization of BRCA1 probes in the two-color FISH technique?
What color indicates the visualization of BRCA1 probes in the two-color FISH technique?
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What type of protein is HER2/neu classified as?
What type of protein is HER2/neu classified as?
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What is the percentage of interphase cells analyzed that were found positive for BCR-ABL gene fusion in the first analysis?
What is the percentage of interphase cells analyzed that were found positive for BCR-ABL gene fusion in the first analysis?
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Which translocation is associated with Acute Promyelocytic Leukemia (APL)?
Which translocation is associated with Acute Promyelocytic Leukemia (APL)?
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What is the primary purpose of the FISH technique mentioned in the content?
What is the primary purpose of the FISH technique mentioned in the content?
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Which medication is noted as a molecularly targeted drug for BCR-ABL positive CML patients?
Which medication is noted as a molecularly targeted drug for BCR-ABL positive CML patients?
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Which chromosomes are involved in the translocation associated with Acute Promyelocytic Leukemia?
Which chromosomes are involved in the translocation associated with Acute Promyelocytic Leukemia?
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What is the expected FISH signal pattern in a normal male cell when hybridized with the CEP X/Y DNA Probe?
What is the expected FISH signal pattern in a normal male cell when hybridized with the CEP X/Y DNA Probe?
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Which of the following genes is located on Chromosome 17 and is associated with susceptibility to breast cancer?
Which of the following genes is located on Chromosome 17 and is associated with susceptibility to breast cancer?
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Which method is NOT mentioned as an FDA-approved technology for breast cancer screening?
Which method is NOT mentioned as an FDA-approved technology for breast cancer screening?
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Study Notes
Fluorescence Microscopy
- Analyzes fluorescence from a sample
- A beam of light (e.g., UV) excites electrons in molecules of certain compounds, causing them to emit light, often visible light
Epifluorescence Microscope
- A type of fluorescence microscope where light passes through the sample in a straight angle, maximizing illumination
- Uses high-energy light (e.g., xenon, mercury lamp), but now LED is used
- Uses fluorescence and phosphorescence
Applications of Fluorescence
- Mineralogy and gemology
- Chemical sensors (fluorescence spectroscopy)
- Fluorescent labeling dyes
- Biological detectors
- Fluorescent lamps
Applications in Cytogenetics
- Routine karyotyping/cytogenetic analysis
- Color karyotyping
- Spectral karyotyping (SKY)
- Multicolor fluorochrome banding (mBand)
- Automatic telomere measurement
Fluorescence in situ Hybridization (FISH)
- A powerful technique used to detect chromosomal abnormalities/mutations
- Introduced in the late 1980s
- Addressed the limitations of routine chromosome analysis
- Refined the level of cytogenetic screening
- High specificity and sensitivity
- Shortened turnaround time (TAT) of patient results
Types of FISH Detection
- Direct FISH: Reporter + Probe
- Indirect FISH: Antibody + Reporter + Probe
Fluorescent in situ Hybridization (FISH) Protocol
- Sample DNA is denatured.
- Probe DNA is denatured and hybridized with the sample DNA.
- Either a direct or indirect label is used.
- The sample is washed and detected.
Different Types of FISH Probes
- Chromosome painting: Targets whole chromosomes
- Centromeric probe: Targets the centromere region
- Telomeric probe: Targets the telomere region
- Gene-specific probe: Targets a specific gene
FISH in Hematological Malignancies
- FISH can be used to identify translocations, deletions, and amplifications in hematological malignancies.
- Example: In Chronic Myelogenous Leukemia (CML), FISH can detect the BCR-ABL fusion gene caused by the t(9;22) translocation.
- Imatinib Mesylate (Gleevec) is a molecularly targeted chemo drug for BCR-ABL positive CML patients.
- Example: In Acute Promyelocytic Leukemia (APL), FISH can detect the PML/RARα gene fusion caused by the t(15;17) translocation.
- Trans Retinoic Acid is a molecularly targeted drug for patients positive for t(15;17)(q24;q21) or PML/RARα gene fusion.
FISH in Bone Marrow Transplantation
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Example: FISH can be used to determine XX/XY cell ratio in sex-mismatched bone marrow transplants using CEP X SpectrumOrange/CEP Y SpectrumGreen DNA Probe Kit:
- In a normal male cell, the expected pattern is one orange, one green (1O1G)
- In a normal female cell, the expected pattern is two orange (2G) signals.
FISH in Solid Tumors
- Example: FISH can be used to detect gene mutations in solid tumors, such as breast cancer and other malignancies.
BRCA1 and BRCA2 Genes
- BRCA1 and BRCA2 are tumor suppressor genes located on chromosomes 13 and 17, respectively.
- Mutations in these genes increase the risk of breast, ovarian, and other cancers.
- Women with abnormal BRCA1 or BRCA2 genes have about a 60% risk of being diagnosed with breast cancer during their lifetimes (compared to 12-13% for women overall).
HER2/neu gene
- HER2/neu (often just HER2) is a growth-promoting protein located on chromosome 17, near the BRCA1 gene locus.
- HER2 gene amplification is associated with aggressive breast cancer.
- Testing for HER2 status can enable the most appropriate therapy decision.
- Tests should measure either gene copy number or the presence of protein receptors.
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Description
Explore the fundamentals of fluorescence microscopy, including epifluorescence techniques and various applications. Learn about its significance in fields such as cytogenetics, mineralogy, and biological detection methods. This quiz will test your understanding of how fluorescence aids in scientific discoveries.
