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Sem 3: Viro > Cultivation of Viruses + Virus Purification & Concentration > Flashcards

Cultivation of Viruses + Virus Purification & Concentration Flashcards

1
Q

What are the possible ways to grow and cultivate viruses?

A
  1. Cell / Tissue culture
  2. Inoculation in embryonated egg
  3. Lab animal inoculation
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2
Q

What are the 2 types of cell culture?

A
  1. Suspension culture
  2. Monolayer culture
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3
Q

What is the mechanism of suspension cultures?

A

cells dont require attachment for growth + dont attach to surface of the culture

  • cells suspended in media / propagated in suspension
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4
Q

What is the mechanism of monolayer cultures?

A

the bottom of the culture is covered with continuous layers of cells, 1 cell in thickness

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5
Q

What are the advantages of primary cell culture?

A
  1. best for isolation + propagation of viruses
  2. Heterogeneous - many cell types
  3. Closest to animal / similar to animal tissue cells
  4. Used to make viral vaccines
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6
Q

What are the disadvantages of primary cell culture?

A
  1. Susceptible to contamination
  2. Complex, may not fully act like parent tissue
  3. Short lifespan in culture
  4. Difficult to obtain
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7
Q

What is subculture?

A

When you transfer cells from primary cell culture to a new cell culture

Ex: primary cell culture to secondary cell culture

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8
Q

Why is subculturing neccessary?

A

Required to provide fresh nutrients and growing space for continuously growing CELL LINES

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9
Q

What types of cell lines are there?

A
  1. Continuous
  2. Finite/ diploid
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10
Q

What is the mechanism of finite/diploid cell lines?

A
  • limited life span with limited # of cell generation

** can only be subcultured a certain amt of times before it cant anymore

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11
Q

What is the mechanism of continuous cell lines?

A

Has the ability to divide indefinitely

** can be continuously subcultured

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12
Q

True or False: In rate-zonal centrifugation, virus particles are separated based on their buoyant density

A

FALSE - based on mass

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13
Q

What ways can be used for purification of viruses?

A
  1. Membrane chromatography
  2. Isopycnic centrifugation
  3. Rate-zonal centrifugation
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14
Q

____________ plays a vital role in virology by efficiently sedimenting the smallest viruses

A

Ultracentrifuge

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15
Q

________ separates viral particles on the basis of their buoyant density and aids in purification and concentration of viruses

A

Isopycnic centrifugation

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16
Q

What is rate-zonal centrifugation?

A

Ultracentrifugation method for purification and concentration of viruses

  • separates virus particles based on their MASS

** bc particles move @ dif rates depending on their mass when under centrifugal force

17
Q

What is a primary cell culture?

A

Maintenance of growth of cells dissociated directly from parental tissue (Ex: kidney or liver) of human or animal origin

18
Q

What are some characteristics of finite/diploid cell lines?

A
  1. Limited life span
  2. Homogenous population of single cell type
  3. Derived from embryos or secondary cell culture
  4. *SLOW GROWTH RATE
  5. Cells retain original morphology
  6. Cells exhibit contact inhibition, density limitations + anchorage dependance
  7. *USED FOR VACCINES!
19
Q

What are the characteristics of immortal/ heteroploid cell lines?

A
  1. Mostly homogenous w single cell type
  2. **DERIVED FROM CANCER CELLS
  3. **FDA PROHIBITS USE FOR VACCINES
  4. Rapid growth rate
  5. No contact inhibition or anchorage dependance
  6. Genetically, furthest away from the animal
20
Q

Which cell line can be used for vaccine production?

A

Finite/ diploid cell lines ONLY

21
Q

Which cell line exhibits anchorage dependance?

A

Finite/diploid

22
Q

What are examples of a continuous cell line?

A
  1. HeLa cells
  2. Henrietta Lacks cells
23
Q

What morphology can viral cells in culture have?

A
  1. Fibroblastic
  2. Epithelial-like
  3. Lymphoblast-like
24
Q

What are the characteristics of a fibroblastic cell in culture?

A
  1. bipolar or multipolar
  2. Elongated shapes
  3. ** grows attached to a substrate
25
Q

What are the characteristics of an epithelial-like cell in culture?

A
  1. polygonal in shape
  2. regular dimensions
  3. **Grows attached to a substrate in discrete patches
26
Q

What are the characteristics of a lymphoblast-like cell in culture?

A
  1. Spherical shape
  2. **Grown in suspension without attaching to a surface
27
Q

What is required in culture media for viruses to grow?

A
  1. AA
  2. Inorganic salts
  3. Vitamins
  4. Glucose
  5. Serum!
28
Q

What kind of serum is used in culture media and is required for growth and maintenance of cells

A

Fetal bovine serum (FBS)

29
Q

Why is serum necessary for culture media?

A

Helps in:
1. Cell adhesion
2. Regulating cell membrane permeability
3. Provides nutrients to cells

30
Q

How can you tell if your culture media has been contaminated?

A

Phenol red pH indicator!

** if it turns from normal red color to a yellow/orange its bad news and means pH is < 7

30
Q

How can the pH of culture media be maintained?

A

By using exogenous CO2

** bc CO2-bicarb based buffer is used

31
Q

How can you prevent contamination of your culture media?

A

Antimicrobial agents!

Prevents mycoplasma, bacT, yeast, molds, etc

32
Q

What proteolytic enzyme is used to detach and dissociate cells while subculturing?

A

Trypsin

** allows transfer of cells to a new cell plate

33
Q

___________ refers to damage or morphological changes to host cells during virus invasion

A

Cytopathic effect or cytopathogenic effect

34
Q

What are the routes of egg inoculation?

A
  1. Yolk sac
  2. Allantoic cavity
  3. Amniotic cavity
  4. Chorioallantoic membrane (CAM)
35
Q

When inoculating an egg with a virus, how can you tell evidence of viral growth?

A
  1. Death of embryo
  2. Paralysis
  3. Stunted growth
  4. Hemorrhage and congestion
  5. Extracellular membrane lesions
  6. Hemagglutins in embryonic fluid
  7. urate deposits in mesonephros
  8. *** POCKS on chorioallantoic membrane (CAM)

Sem 3: Viro (17 decks)

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