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Biol 466 > Final review-Methods > Flashcards

Final review-Methods Flashcards

1
Q

Flow Cytometry

A

used to measure size, shape, complexities of cells
-can observe surface protein/glycoprotein expression

Adv:
-accurate cell sorting, subpop analysis

Dis:
-too slow, commonly unnecessary information for single experiment

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2
Q

GFP

A

Used to tag specific proteins

Adv:

  • analyze live organisms
  • study viral infections (pre and post infection)
  • can study across multiple time points

Dis:
-fluorescent protein may interfere with biology

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3
Q

Southern blot

A 
Uses probe (difference to PCR)
-determine DNA fragment size
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4
Q

Elisa

A

Measure concentration of antibody OR antigen

Assay diagram:
measuring antibody:
-antigen-antibody of interest-labeled antibody

measuring antigen:
-antibody-antigen of interest-antibody-labeled antibody

ADV:
-highly sensitive and specific

DIS:
-labeling can effect immunoreactivity of antibody

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5
Q

Northern blot

A

Measure RNA levels

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6
Q

microarray

A

Measure RNA levels

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7
Q

In-situ hybridization

A

determine RNA or DNA presence in tissue

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8
Q

knock-out

A

Removing a gene

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9
Q

knock-down

A

silencing a gene at mRNA level (study function of gene expression without removing it)

  • easy gene rescue
  • gene not completely removed
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10
Q

Animal models

A

used in correlation & causation studies

Adv:

  • use KO studies with controls possible
  • can use clonally identical animals

Dis:

  • some ethical concerns
  • still only models (not humans)
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11
Q

Human models

A

Used usually in correlational studies

Adv:
-best representation of human biology

Dis:

  • control studies not always possible
  • lethal studies not possible
  • ethical concerns
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12
Q

gene trapping

A

randomly mutating genes to try to find gene of interest

ADV:
-wide approach (used when unsure where to start)

DIS:
-non-specific

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13
Q

microarray

A

removing mRNA to determine expression
–>used to determine what mRNA are expressed in sample

ADV:
-massive parallel sequencing

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14
Q

Humanized animal models

A

inject animal with human cells-mimics physiology of disease in humans

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15
Q

cell-line

A
  • cells replicate indefinitely
  • used for long experiments (over 2 weeks)
  • study cancer
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16
Q

CRISPR/CAS9

A

Cuts a gene at a specific location and can insert another-effects DNA (transcription)
-create loss of function OR gain of function mutations

ADV:
-specifically targeted locations on DNA

DIS:
-possible unwanted modifications due to host repair system

17
Q

shRNA

A

creates hairpin at gene of choice-effects RNA (translation

18
Q

organoids

A
  • observe cell-cell interactions

- observe viral infections in organ systems

19
Q

cultures with 2 cell types

A

can mimic more of physiology of disease in humans compared to one cell type cultures
-reflects real cell interactions

  • study cancer
  • study tissue invasion
20
Q

gene rescue

A

Method to see if WT phenotype is restored

Dis: assumes gene was removed and then re-introduced

21
Q

Western Blot

A

Used to detect protein and divide them by size
(denatures the proteins)
-measure antigen concentrations

ASSAY diagram:

  • smallest
  • medium sized
22
Q

Elispot

A

Measure amount of antigen or antibody
-monitor cell immune responses

ASSAY diagram:

  • all 3
  • largest measures antibody
  • other 2 measures antigen

ADV:
-sensitive, allows for detection of single cell

DIS:
-cannot quantify amount of protein secreted by cell

23
Q

immunohistochemsitry

A

Staining of tissue sections to see localization and distribution of specific protein

ADV:
-can view localization of proteins under microscope

DIS:
-fixed samples, cannot observe cellular functioning

24
Q

Plaque assay

A

measures viral concentration based on infection dose

  • used to purify clonal population of virus
  • or determine quantity of infectious virus
  • cell lysis = viral presence

ADV:
-quantitative: determine viral titres

DIS:
-some viruses don’t cause cell lysis

25
Q

FFA (focus forming assay)

A

variation of plaque assay

-detects infected host cells and infectious viral particles

26
Q

Passage

A

process of growing bacteria/virus in iterations

-can study changes in virulence of pathogen (over time)

27
Q

induced pluripotent stem cells

A

observe viral infections in organ system

observe cell-cell interaction in patient sample

28
Q

primary cell culture

A

comes from real patient sample of their cells

29
Q

confocal microscopy

A

magnification of cells under microscope

-often used with fluorescence

Biol 466 (3 decks)

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