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CYTOGENETIC TECHNIQUES Flashcards

1
Q
  • Study of the physical size and structure of chromosomes
  • A specialized laboratory discipline that examine the structure and behavior of chromosomes at the cellular level
A

Cytogenetics

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2
Q

cells are arrested in what phase of mitosis?

A

end prophase or in early
metaphase

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3
Q

cells are arrested in end prophase or in early metaphase of mitosis because of

A
  • Compact and densely staining of chromosomes
  • Characteristic size and shape of chromosomes
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4
Q
  • Checks the number of chromosomes in the normal diploid cell
  • Check the size distribution
A

Karyotype

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5
Q

Comparing chromosomes are based on:

A
  • Length
  • Location of centromeres
  • Location and sizes of G-bands
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6
Q

Two Cytogenetic Techniques

A
  1. Classical or Standard Cytogenetic
  2. Fluorescence in situ hybridization (FISH)
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7
Q

Allows visualization of loss or gain of material

A

Classical or Standard Cytogenetic

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8
Q
  • Allows much smaller changes to be seen
  • Far more advanced
A

Fluorescence in situ hybridization (FISH)

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9
Q

Blood additives

A

EDTA, ACD

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10
Q

what is EDTA?

A

EthyleneDiamineTetraAcetic Acid

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11
Q
  • Prevents the coagulation of blood
  • Allows the qualitative and quantitative determination of HIV, Hepatitis C and Cytomegalovirus (CMV)
A

EDTA

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12
Q
  • Prevents the coagulation of blood
  • Preserve the form and function of cellular components
A

Acid Citrate Dextrose

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13
Q

top color of EDTA?

A

purple top

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14
Q

top color of ACD?

A

yellow top

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15
Q
  • Uncommon
  • Used for:
  • Bladder cancer screening
  • Monitor therapy of bladder cancer
A

Urine

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16
Q

type of urine used

A

midstream

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17
Q

how much urine is needed?

A

first 10ml

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18
Q
  • Differential and quantitative analysis as a sensitive specific biomarker for the detection of colorectal cancer
A

Feces

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19
Q
  • Rapid identification of infectious agents
  • Comes from brain and spinal cord
A

Cerebrospinal Fluid

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20
Q

how many tubes are used in CSF? what tube is used?

A

4 tubes are collected; 2nd tube is used

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21
Q

why is the 2nd tube used for CSF collection?

A

it is the purest CSF specimen

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22
Q

normal CSF protein

A

50-80 mg/dL
0.18-0.45 g/L

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23
Q

normal CSF glucose

A

15-45 mg/dL
2.5-3.5 mmol/L

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24
Q

normal CSF RBC count

A

0 cells/mm3

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25
Q

normal CSF WBC count

A

0-3 cells/mm3

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26
Q

normal CSF xanthochromia (yellowish color)

A

none

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27
Q

normal CSF appearance

A

normal (water)

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28
Q

bacterial CSF appearance

A

turbid

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29
Q

viral CSF appearance

A

clear

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30
Q

fungal CSF appearance

A

fibrin web (clotting)

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31
Q

bacterial CSF protein

A

more than 1

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32
Q

viral CSF protein

A

less than 1

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33
Q

fungal CSF protein

A

0.1-0.5

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34
Q

bacterial CSF glucose

A

less than 2.2 mmol/L

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35
Q

viral CSF glucose

A

normal (2.5-3.5)

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36
Q

fungal CSF glucose

A

1.6-2.5

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37
Q

bacterial CSF WBC

A

more than 500

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38
Q

viral CSF WBC

A

less than 1000

39
Q

fungal CSF WBC

40
Q

bacterial CSF increased in what type of WBC?

A

neutrophils

41
Q

viral and fungal CSF increased in what type of WBC?

42
Q
  • Assess the presence of infectious microorganisms
  • Fluid found in joints
A

synovial fluid

43
Q

process of collecting fluids from the joints

A

arthrocentesis

44
Q

tube used in arthrocentesis?

A

Sterile plain tube

45
Q

clarity, color, and viscosity of normal SVF (synovial fluid)

A

transparent, clear, high

46
Q

clarity, color, and viscosity of non-inflammatory SVF (synovial fluid)

A

transparent, yellow, high

47
Q

clarity, color, and viscosity of inflammatory SVF (synovial fluid)

A

translucent, yellow, low

48
Q

clarity, color, and viscosity of septic SVF (synovial fluid)

A

opaque, dirty/yellow, variable

49
Q

clarity, color, and viscosity of hemorrhagic SVF (synovial fluid)

A

bloody, red, variable

50
Q

wbc count and PMNs% of normal SF

A

less than 200, less than 25%

51
Q

wbc count and PMNs% of non-inflammatory SF

A

200-2000, less than 25%

52
Q

wbc count and PMNs% of inflammatory SF

A

200-10000 (up to 100000), more than 50%

53
Q

wbc count and PMNs% of septic SF

A

more than 80,000, more than 75%

54
Q

wbc count and PMNs% of hemorrhagic SF

A

200-2000, 50-75%

55
Q
  • Prenatal diagnosis of congenital disorder
  • Assess fetal maturity
  • To look for Rh isoimmunization or intrauterine infection (blood type infections)
A

amniotic fluid

56
Q

when to collect amniotic fluid?

A

15-20 weeks AOG (4th-5th month)

57
Q

how many days does amniotic fluid culture take?

58
Q
  • For earlier diagnosis than amniotic fluid
  • part of the placenta
A

chorionic villus

59
Q

when to collect chorionic villus?

A

10-12 weeks (2nd-3rd month)

60
Q

how many days does chorionic villus culture take?

61
Q
  • For infectious agents identification
  • Identification of cancer cells
A

Pleural, pericardial, ascitic fluid

62
Q

collection of fluid from the lungs, heart, and abdomen

A

Paracentesis

63
Q
  • Less invasive
  • Excellent source of genomic DNA
  • For patients who had blood transfusion
    and bone marrow transplantation
A

Buccal cells

64
Q

method of collecting buccal cells

A
  • Rinsing with mouthwash
  • Use of swab
65
Q
  • Formalin-fixed, paraffinembedded tissue
  • Frozen specimen in an Optimal Cutting Temperature Compound (OCT)
A

solid tissue

66
Q
  • Forensic analysis (Genomic DNA Identification)
  • Trace metal and drug analysis
A

Hair and nails

67
Q

specimens transported in room temp

A
  • blood
  • bone marrow
  • amniotic fluid
  • chorionic villi
68
Q

specimens transported in ice

A

solid tissue

69
Q

Techniques in Classical Cytogenetics

A
  1. Cell culture
  2. Sufficient metaphase plates are produced
  3. Cells are subjected to hypotonic swelling
  4. Fixation
  5. Staining
  6. Photomicroscopy
  7. Case report
70
Q

multiplication of cells using mitogenic stimulating agents

A

cell culture

71
Q

chemical agent which stimulates mitosis?

A

phytohemagglutinin (PHA)

72
Q

arresting the cells at end prophase using mitotic inhibitors

A

Sufficient metaphase plates are produced

73
Q

mitotic inhibitor used

74
Q

To further disperse the chromosome within the cell and to lyse any red cells present (to harvest the chromosomes)

A

hypotonic swelling

75
Q

buffer solution used in hypotonic swelling

A

0.075M KCl

76
Q

how long are cells stored in hypotonic swelling?

A

10-20 mins, 37 deg Celsius

77
Q

Alters the cell membranes and chromosomes by removing lipids and water molecules and denature proteins

78
Q

modified Carnoy’s fixative

A

3:1
absolute methanol : glacial acetic acid

79
Q
  • Artificial aging of the cells at 65C for 30-60 mins improves the quality of the staining
  • Dehydrate the chromosome
80
Q

dehydration of chromosomes are treated with?

A

proteolytic enzyme
solution

81
Q

what dye is used in staining?

A

metachromatic dye

82
Q

Lightly stained

A

Euchromatin

83
Q

Dark stained

A

Heterochromatin

84
Q

Use computer imaging systems with software to capture and karyotype 2 or more representative metaphase spreads per specimen

A

Photomicroscopy

85
Q
  • Description of the chromosomal finding in International System of Cytogenic Nomenclature (ISCN)
  • Recommendations are made
    – When abnormalities are detected
    – Genetic counseling
    – Referral to specialist
    – Additional genetic testing
A

Case report

86
Q
  • Uses fluorescent dye to mark abnormalities
  • The principal molecular technique currently used in clinical cytogenetic laboratory
  • Called Molecular Cytogenetics
  • Involves applying DNA probes to a chromosome spread
A

Fluorescent In Situ Hybridization

87
Q

Process of FISH

A
  1. Chromosomes are placed on a microscope slide and then denatured
  2. Complimentary DNA probes are constructed
  3. When the DNA is denatured, the probe can then hybridize (form hydrogen bonds) to the complementary single-stranded DNA
  4. The labeled probes can be used to identify different chromosomes or targeted chromosome regions
88
Q

separates the ouble helix into single strands using heat, chemicals and pH change

A

Denaturing

89
Q
  • For the sequence of interest and labeled with a fluorescent dye.
  • Ultraviolet microscope
    (fluorescence microscope)
A

Complimentary DNA probes are constructed

90
Q

the things that light up under the microscope

A

fluorophores

91
Q
  • Detected using a fluorescence microscope
  • The probe will be visible as one or more fluorescent signals in the microscope
A

When the DNA is denatured, the probe can then hybridize (form hydrogen bonds) to the complementary single-stranded DNA

92
Q

To help recognize:
– Trisomies
– Translocations
– Deletions

A

The labeled probes can be used to identify different chromosomes or targeted chromosome regions

93
Q

Uses of FISH

A
  • To form a diagnosis
  • Evaluate a prognosis
  • Evaluate remission of a disease
    – Cancer
94
Q

FISH vs Classical Cytogenetics

A
  • Detect diseased cells more easily
  • Does not require living cells
  • Can be quantified automatically
  • A computer counts the fluorescent dots present

CYTOGENETICS (11 decks)

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