Lec 3: Methods Flashcards

1
Q

methods in virology are used also in

A

cell & molecular biology

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2
Q

most cells taken from body…

A

don’t grow well in culture

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3
Q

cell line =

A

If cells from a primary culture can be subcultured they are growing as a cell line

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4
Q

continuous cell line =

A

cells can be subcultured only a finite number of times unless they are immortalized, which can be subcultured indefinitely as a continuous cell line

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5
Q

cancer cells are already…

&…

A

immortalized,

…& continuous cell lines may be established from these without further treatment

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6
Q

Normal Cells from Animal –> _____ –> —-(___)—-> ____ —-(______)—-> _____

A

Normal Cells from Animal –> 1* Cell Culture –> —-(subculture)—-> Cell Line —-(immortalization)—-> Continuous Cell Line

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7
Q

Cancer Cells from Animal –> _____

A

Cancer Cells from Animal –> Continuous Cell Line

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8
Q

primary culture cells have

A

finite life span

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9
Q

Primary culture contains

A

a very heterogeneous population of cells

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10
Q

Subculturing of primary cells leads to

A

the generation of cell lines

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11
Q

Cell lines have

A

limited life span, they passage several times before they become senescent

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12
Q

Cells such as macrophages and neurons do not

A

divide in vitro so can not be used as primary cultures

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13
Q

cell strain

A

Lineage of cells origination from the primary culture is called a cell strain

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14
Q

Continuous Cell Lines –>

A

HeLa cells

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15
Q

senescence =

A

Normal cells usually divide only a limited number of times before losing their ability to proliferate, which is a genetically determined event known as senescence; these cell lines are known as finite.

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16
Q

Some cell lines become…
through a process called…
which can…

A

…become immortal,
…through transformation,
…can occur spontaneously or can be chemically or virally induced

17
Q

continuous cell lines form when

A

a finite cell line undergoes transformation and acquires the ability to divide indefinitely

18
Q

continuous cell lines have undergone significant…

this can…

A

…mutations to become immortal,

…can alter the biology of the cell and must be taken into consideration in any analysis.

19
Q

Why do you need agar in the media? (2)

A
  • To keep virus particles in place, otherwise they would disperse everywhere
  • Allows you to control movement of the virus
20
Q

(after centrifugation)
top band =
bottom band =
each band =

A

lowest density
highest density
pure sample of virus

21
Q

viral structure studies uses

A

microscopy:

  • light = not suitable due to limited resolution
  • electron = negative staining (stain w/ heavy metal; stained portion will reflect the electron beam) (heavy metal = white dot ) (no metal/structure = black dot)
22
Q

x-ray beam array =

A

viral protein analysis
allows for diffraction pattern determination and relative positions of molecules to be determined
(important for drug design)

23
Q

Detection of: Virions –>

A

Electron Microscopy

24
Q

Detection of: Infectivity of cell culture –>

A

Examine change in characteristics of the cells (plaque arrays?)

25
Q

Detection of: Virus Antigens –>

A

ELISA

26
Q

Detection of: Virus Antibodies –>

A

ELISA

27
Q

Detection of: Virus Nucleic Acids –>

A

Hybridization (southern, northern) & PCR

28
Q

SDS =

this will make…

A

“Sodium Dodecyl Sulfate”
breaks bonds in proteins, making them all simple 1* chains
…this will make proteins move through electrophoresis gel at a rate dependent only on MW

29
Q

PAGE =

A

Polyacrylamide gel electrophoresis

30
Q

use PAGE for

A

proteins

31
Q

PAGE makes proteins move from

A

cathode (-) —> anode (+)

32
Q

Principles of tests to detect virus antigens:

A

Direct Test = anti-virus antibody labelled

Indirect Test = second antibody labelled

33
Q

Blotting and probing for detection of:

  • DNA =
  • RNA =
  • Proteins =
A

= southern blot/ hybridization
= northern blot/hybridization)
= western blot

34
Q

PCR aka

A

Polymerase chain reaction

35
Q

(infectivity assays)

eclipse period =

A

period during which no intracellular infectious virus can be recovered;
- infectious nucleic acid might
be recoverable in some cases

36
Q

(infectivity assays)

bust size

A

average yield of infectious virus per cell