lab techniques Flashcards

(24 cards)

1
Q

what are immune mediators

A
  • antigens
  • antibodies
  • cytokines
  • cells
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2
Q

what does the concentration of immune mediators tell us

A

information of the disease state - mechanism, diagnosis, response to treatment

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3
Q

what occurs with a blood group assay

A
  • dilute blood sample
  • add antibodies against erythrocyte antigens (A, B ,Rh)
  • agglutination occurs to determine blood group
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4
Q

what does a radioimmunoassay do

A
  • competitive antibody binding assay
  • cold insulin competes with hot insulin
  • higher 125 I insulin means less normal insulin and type 1 diabetes
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5
Q

what is the disadvantage of radioimmunoassay

A

radiolabel is hazardous

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6
Q

what is the advantage of radioimmunoassay

A

faster than chromatography based methods

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7
Q

what does a low sample of insulin present as in an radioimmunoassay

A
  • low conc of 125I labelled insulin
  • high radiolabelled signal
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8
Q

what does a high sample of insulin present like in an radioimmunoassay

A
  • high conc of 125 I labelled insulin
  • low radiolabelled signal
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9
Q

what does ELISA stand for

A

enzyme linked immunosorbent assay

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10
Q

what are the types of ELISA

A

direct, indirect, competitive and non-competitive

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11
Q

what does an indirect, non competitive ELISA do

A
  • measures specific antibody concentration
  • antigen adsorbed to well surface
  • blocking solution blocks non-specific binding
  • add primary antibody in excess and wash off unbound
  • add conjugated antibody in excess
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12
Q

what is a conjugate in ELISA

A
  • an enzyme - catalyses further reaction
  • florescent molecule - spectrophotometry
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13
Q

how is an indirect, non competitive ELISA used in clinical practice

A
  • detection of antibodies in response to infection
  • diagnostic for HIV
  • detection of antibodies for immunisation eg. Hep B
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14
Q

why use a indirect ELISA over direct

A
  • every antibody labelled individually
  • saves time, labour costs and increases safety
  • increases sensitivity
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15
Q

how do you use ELISA to measure antigen concentration

A
  • antigen filling between two antibodies (sandwich)
  • first captures with a high affinity and next recognises epitope
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16
Q

what does western blotting do

A
  • proteins have a + and - charge
  • separates protein sample on SDS-polyacrylamide gel by electrophoresis
17
Q

what does western blotting tell us

A
  • distance travelled - estimation of molecular weight
  • amount of chemiluminescent signal allows estimation of abundance
18
Q

how is western blotting used in clinical practice

A
  • conformation test for anti-HIV antibodies
19
Q

what are rapid/ point of care tests used for

A
  • detection of antibodies for HIV, hep C, covid-19
  • detection of antigen for malaria, ebola, influenza, strep, HCG
20
Q

what does immunofluorescence show

A

visualisation of tissue, structures, cells and sub- cellular components

21
Q

how does immunofluorescence work

A
  • need to be able to detect if antibodies react with cellular antigens
22
Q

what does flow cytometry do

A

Counts cells based on size,
contents, and cell surface
expression

23
Q

what are the steps in flow cytomerty

A

1- prepare single cell suspension
2- make fluorescently labelled antibodies
3- stain patients leukocytes
4- count on Flow cytometer