FISH Flashcards
1
Q
FISH uses ___ light
A
excitation
2
Q
A cytogenetic technique that uses fluorescent probes that bind specifically to a part of chromosomes complimentary to its sequence
A
FISH
3
Q
Small strips of single stranded DNA complimentary to the sites we want to examine
A
Probes
4
Q
Probes are ___ (natural/artificial) DNA
A
artificial
5
Q
Target chromosomes are ___ (natural/artificial) DNA
A
natural
6
Q
Useful in detecting and mapping the presence or absence of particular DNA sequence within chromosomes
A
FISH
7
Q
Number of kb used in FISH
A
200-400
8
Q
Applied to provide specific localization of genes on chromosomes
A
FISH
9
Q
FISH is also used for the rapid diagnosis of ___ and ___ which is acquired using specific probes
A
trisomies
microdeletions
10
Q
FISH is ___ (more/less) cost effective than PCR
A
more
11
Q
FISH has ___ (short/long) turn around time
A
short
12
Q
The basic requirement for FISH is for the DNA to be ___/___/___
A
undenatured
preserved
intact
13
Q
The target DNA is also called as the ___
A
template
14
Q
To separate DNA strands and allow probe access to target DNA
A
Denature
15
Q
Binding the probe to target DNA
A
Hybridize
16
Q
Specimens used for metaphase cell FISH
A
Blood
Bone Marrow
Skin Fibroblast
Chorionic villi
Amniocytes
Tumors
17
Q
Specimens used for interphase FISH
Direct preparation
A
Uncultured cells from blood, bone marrow, cytospins
Smears made from blood, buccal cells, bone marrow
18
Q
Specimens used for interphase FISH
Indirect preparation (Paraffin Embedded Tissue Sections)
A
Tumors
Products of conceptions
19
Q
FFPE means ___
A
Formalin fixed paraffin embedded tissue
20
Q
Metaphase/Interphase FISH:
Gold standard and routinely done
A
Metaphase
21
Q
Metaphase/Interphase FISH:
Done on cultured cells
A
Metaphase
22
Q
Metaphase/Interphase FISH:
Allows direct visualization of chromosomes and exact position of signals
A
Metaphase
23
Q
Metaphase/Interphase FISH:
Useful in the detection of structural changes in the genokme
A
Metaphase
24
Q
Metaphase/Interphase FISH:
More rigorous procedure
A
Metaphase
25
Metaphase/Interphase FISH:
Used for copy number changes
Metaphase
26
Metaphase/Interphase FISH:
Can be modified for de novo detection
Metaphase
27
Metaphase/Interphase FISH:
May also be done on uncultured specimens
Interphase
28
Metaphase/Interphase FISH:
Advantageous in the rapid screening of many nuclei for prenatal diagnosis and newborn studies
Interphase
29
Metaphase/Interphase FISH:
Also beneficial in the study of samples with a low mitotic index such as most solid tumors
Interphase
30
Metaphase/Interphase FISH:
Major disadvantage is the inability to detect unknown structural chromosomal changes
Interphase
31
Metaphase/Interphase FISH:
Same day turn around time
Interphase
32
Metaphase/Interphase FISH:
Translocation can be detected
Interphase
33
Metaphase/Interphase FISH:
Used for screening
Interphase
34
Sample for interphase fish:
For ploidy analysis during prenatal studies
Amniocytes
35
Sample for interphase fish:
For ploidy analysis in newborns
Peripheral blood smears
36
Sample for interphase fish:
Translocation or copy number analysis in cancer studies
Bone marrow aspirate smear or direct harvest
37
Complementary sequences of target nucleic acids tagged or labeled with fluorophores
FISH probes
38
FISH probes can be (3):
DNA
RNA
Nucleic acid analogs
39
FISH probes size ranges from __ to __ base pairs
20-1000
40
Direct/Indirect labeling:
Fluorophores are directly attached to the probe
Direct
41
Direct/Indirect labeling:
Less sensitive
Direct
42
Direct/Indirect labeling:
FITC, rhodamine, cyanines
Direct
43
Direct/Indirect labeling:
Chemical conjugation of the nucleic acid with a nonfluorescent molecule that can bind fluorescent material after hybridization
Indirect
44
Direct/Indirect labeling:
Biotin & digoxigenin
Indirect
45
Partner molecule of biotin
Avidin
46
A process of sticking DNA back to its original double stranded structure which requires the lowering of temperature
Reannealing
47
This is usually washed off that can cause background fluorescence/"noise" and lessens sensitivity
Excess unbound probe
48
Allows the visualization of nuclear boundaries and morphology
counterstain
49
Examples of counterstain
Propidium iodide
DAPI
50
T/F: Repetitive target does not brighten the fluorescence because they are often small
F (may brighten fluorescence even with small prove because of repetitive sequence)
51
In denaturing the DNA, ___ and ___ are used
heat
denaturing chemical (formamide/salt solution)
52
Most commonly used FISH probes (3)
Whole chromosome paints (WCPs)
Chromosome enumeration probes (CEPs)
Locus-specific identifiers (LSIs)
53
Most commonly used FISH probe:
Hybridize to the unique sequences which cover the length of an entire chromosome or chromosome arm
WCP
54
Most commonly used FISH probe:
Useful in studying marker chromosomes, translocations, and aneuploidy in metaphase
WCP
55
Most commonly used FISH probe:
NOT useful in interphase cells
WCP
56
Most commonly used FISH probe:
Collection of smaller probes that bind to the whole length of chromosome
WCP
57
Most commonly used FISH probe:
Useful in the examination of chromosomal aberrations
WCP
58
Most commonly used FISH probe:
Also known as alpha-satellite probes as they bind to highly repetitive DNA
CEP
59
Most commonly used FISH probe:
Applicable for both metaphase & interphase FISH
CEP
60
Most commonly used FISH probe:
Frequently hybridized to centromere regions
CEP
61
Most commonly used FISH probe:
Most often used as control probes
CEP
62
Most commonly used FISH probe:
Most probes give large, bright signal
CEP
63
Most commonly used FISH probe:
Useful for detecting aneuploidy
CEP
64
Most commonly used FISH probe:
Generated from repetitive sequences found in the middle of each chromosome
Alphoid/Centromeric repeat
65
Most commonly used FISH probe:
Used to determine whether an individual has the correct number of chromosomes or if there is aneuploidy in the patient's genome
Alphoid/Centromeric repeat
66
Most commonly used FISH probe:
More gene/region specific
LSI
67
Most commonly used FISH probe:
Hybridize to unique sequences in the genome
LSI
68
Most commonly used FISH probe:
Probes span the specific region and average 200-300 kb in size
LSI
69
Most commonly used FISH probe:
Useful diagnostic tool to detect deletions, duplications, rearrangements, amplifications
LSI
70
Most commonly used FISH probe:
Can also be used in fusion-type probe strategies
LSI
71
Most commonly used FISH probe:
Binds to a particular regions of a chromosome
LSI
72
Most commonly used FISH probe:
Used when only a small portion of a gene is isolated and want to determine on which chromosome the gene is located, or how many copies of a gene exist within a particular genome
LSI
73
Specialize and evolving technologies:
A technique that allows the investigator to view a karyotype so that each chromosome is "painted" with a different color
Multiplex FISH (M-FISH)
74
Specialize and evolving technologies:
Ratio-labeled probes are used to create a distinct computer-generated false color for each chromosome
Multiplex FISH (M-FISH)
75
Specialize and evolving technologies:
Useful for complex rearrangements, such as those seen in neoplastic disorders and solid tumors
Multiplex FISH (M-FISH)
76
Specialize and evolving technologies:
Uses chromosome-specific mixtures of partial chromosome paints that are labeled with various fluorochromes
Multicolor Banding (mBAND) Analysis
77
Specialize and evolving technologies:
A computer program analyzes metaphase chromosome data and procedures a pseudocolored banded karyotype with an estimated resolution of 550 bands, regardless of chromosome length
Multicolor Banding (mBAND) Analysis
78
Specialize and evolving technologies:
Advantageous for the determination of breakpoints and the analysis of intrachromomal rearrangements and can be particularly useful in preparations with shorter chromosomes
Multicolor Banding (mBAND) Analysis
79
Single/Dual color FISH probe:
Designed to cover a gene of interest
Single
80
Single/Dual color FISH probe:
Allows simultaneous detection of numerical abnormalities of two to three regions ins one FISH assay
Dual
81
Probe strategies:
Probes span specific regions of interest, typically regions with recurring breakpoints
Dual fusion FISH probe strategies
82
Probe strategies:
Used to identify specific chromosome rearrangements, typically involving 2 chromosomes
Dual fusion FISH probe strategies
83
Probe strategies:
Probes will flank the specific region of interest
Break apart (BAP) FISH probe strategies
84
Probe strategies:
Used to detect chromosomal rearrangements
Break apart (BAP) FISH probe strategies
85
Probe strategies:
Useful in FFPE sample
Break apart (BAP) FISH probe strategies
86
Probe strategies:
Useful for rearrangements where there are multiple known partners
Break apart (BAP) FISH probe strategies
87
Probe strategies:
Applicable for knowing the source of translocation material
Break apart (BAP) FISH probe strategies
88
Types of FISH probes:
Specific to the subtelomere region of chromosome
Subtelomere
89
Types of FISH probes:
Useful in the detection of subtelomere deletions and rearrangements
Subtelomere
90
Types of FISH probes:
Comprise of different combinations of fluorophore-labeled probes specific for chromosomes 13, 18, 21, X, and/or Y
Pre-Natal
91
T/F:
Cell scoring in FISH: If there are two nuclei with one signal each or one twisted nuclei, do not count
T
92
T/F:
Cell scoring in FISH: If there is one diffused and one clustered signal, do not count
F (must be counted as two signals)
93
T/F:
Cell scoring in FISH: If there is one normal and one split signal, count as two signals
T
94
Specialized and evolving technologies:
A technique that is almost entirely used for research
Fiber FISH
95
Specialized and evolving technologies:
Allows the chromosomes to be stretched out and elongated
Fiber FISH
96
Specialized and evolving technologies:
Provides a much higher spatial resolution and allows for correct orientation and placement of probes and for precise mapping of proves
Fiber FISH
97
Specialized and evolving technologies:
Essentially PCR on a slide
Primed In Situ Labeling (PRINS)
98
Specialized and evolving technologies:
Primers of interest are hybridized on a slide and then subjected to cycles of denaturation, reannealing, and elongation that are used to incorporate labeled nucleotides
Primed In Situ Labeling (PRINS)
99
Specialized and evolving technologies:
Can differentiate hybridizaton with the alpha satellite sequences for chromosomes 13 and 21, something that cannot be done with traditional FISH
Primed In Situ Labeling (PRINS)
100
Specialized and evolving technologies:
Used to identify material of unknown origin
Reverse FISH
101
Specialized and evolving technologies:
A technique that uses DNA from the cells of interest, rather than using a standard karyotype, for chromosomal analysis
CGH
102
Specialized and evolving technologies:
Can be very useful, especially in some cancers when only DNA is available rather than any growing cells
CGH
103
Specialized and evolving technologies:
Used successfully for clinical analysis, particularly with cases that have a low (or no) mitotic index
CGH
104
Specialized and evolving technologies:
Not useful for detecting balanced rearrangements
CGH
