3. lab diagnostics

Question 1

pathology

Answer 1

study of disease

Question 2

aetiology

Answer 2

causation

Question 3

pathogenesis

Answer 3

progression of disease leading to effects on cells, tissues, and organs

Question 4

potential causes of disease

Answer 4

• pathogenic organisms
• genetic mutations
• epigenetic abnormalities
• nutrient deficiencies
• harmful environmental stimuli

Question 5

molecular manifestations of disease

Answer 5

changes in protein
- expression
- structure
- function

Question 6

cellular manifestations of disease

Answer 6

changes in cell
- number
- morphology

Question 7

tissue/organ manifestations of disease

Answer 7

gross changes in
- structure
- function

Question 8

BCR-ABL kinase

Answer 8

novel protein
- promotes cell proliferation
- impairs DNA repair
- induces genomic instability
causes >90% of chronic myeloid leukaemia cases but found in other cancers too

Question 9

aetiology of BCR-ABL kinase expression

Answer 9

chromosomal translocation between chromosomes 9 and 22 forming BCR-ABL fusion gene

Question 10

pathogenesis of BCR-ABL kinase

Answer 10

• hyperplasia
• dysplasia
• neoplasia

Question 11

diagnostic techniques investigate

Answer 11

potential changes associated with
- aetiology
- pathogenesis

Question 12

what is something that halts pcr extension

Answer 12

extension cannot continue through a primer

Question 13

DNA polymerase PCR cofactor

Answer 13

magnesium chloride

Question 14

BCR-ABL treatment

Answer 14

targeted drug therapy

Question 15

BCR-ABL test

Answer 15

BCR pcr - control for primers
BCR-ABL pcr - test for fusion gene

Question 16

what is the importance of controls

Answer 16

to ensure all protocols are functioning appropriately to give an accurate result

Question 17

importance of non-template control

Answer 17

to gauge whether contamination has taken place

Question 18

benefits of PCR multiplex to test for BCR-ABL

Answer 18

the BCR PCR can be used as a negative control

Question 19

what would a negative BCR-ABL test mean for a leukaemia patient

Answer 19

the BCR-ABL gene is not responsible for their leukaemia

Question 20

why are enzymes used for diagnostic IHC

Answer 20

they often don’t have the required microscope for fluorescent tag visualisation

Question 21

benefits of fluorescent tags for IHC

Answer 21

can put lots of tags on

Question 22

key stages of IHC

Answer 22

• antigen retrieval
• endogenous peroxidase deactivation
• non-specific blocking
• primary antibody
• secondary antibody
• substrate

Question 23

purpose of IHC

Answer 23

identify the expression of specific proteins in a tissue sample

Question 24

what principle does IHC rely on

Answer 24

all proteins are potential antigens detectable to the immune system so it is possible to generate an antibody that will bind to specific proteins.

Question 25

how does detection work in IHC

Answer 25

the detection antibody often has an enzyme attached that can produce coloured product from substrate. this stains cells that are positive for target proteins.

Question 26

CK7

Answer 26

cytokeratin 7 often expressed in adenocarcinomas of the breast

Question 27

CK20

Answer 27

cytokeratin 20 most often present in colorectal cancer

Question 28

IHC antigen retrieval

Answer 28

tissue sections are heated in a controlled environment to make sure that the antigens in the tissue are exposed for antibodies to bind to

Question 29

IHC Endogenous peroxidase deactivation

Answer 29

blocking or impairment of endogenous peroxidases present in tissue to prevent them from converting the substrate into a coloured product

Question 30

IHC non-specific blocking

Answer 30

non-specific proteins that may bind antigen-specific antibody are blocked, often with bovine serum albumin (BSA) or fetal calf serum (FCS)

Question 31

IHC primary antibody

Answer 31

the antigen-specific antibody is added to the tissue and binds to any antigens that are present. unbound primary antibody is washed away

Question 32

IHC secondary antibody

Answer 32

this is a detection antibody specific for the primary antibody, and often has an enzyme conjugated to it such as horseradish peroxidase (HRP). unbound is washed away

Question 33

IHC substrate

Answer 33

a solution containing hydrogen peroxide and DAB chromagen is added. in the presence of HRP, these reagents produce a coloured product that will stain cells