Molecular Diagnostics of Cancer 3

Question 1

What are biomarkers in cancer?

Answer 1

Biomarkers are produced by cancer cells or by the body in response to cancer.

Question 2

Where can biomarkers be found?

Answer 2

Biomarkers can be found in blood, urine, stool, and biopsies.

Question 3

What are the uses of biomarkers?

Answer 3

Biomarkers are used for diagnosis, prognosis, and treatment monitoring.

Question 4

What is immunodetection?

Answer 4

Immunodetection is the detection of a specific antigen using antigen binding.

Question 5

What is ELISA?

Answer 5

ELISA (Enzyme-Linked Immunosorbent Assay) detects specific antigens/antibodies in a sample.

Question 6

What is the purpose of ELISA?

Answer 6

ELISA is useful in research and clinically to detect infection or disease markers.

Question 7

What do enzyme-linked secondary antibodies do in ELISA?

Answer 7

Enzyme-linked secondary antibodies amplify the signal in the detection process.

Question 8

What is the role of the detection antibody in ELISA?

Answer 8

The detection antibody binds to the protein of interest if it is present in the sample.

Question 9

What happens after adding the detection antibody in ELISA?

Answer 9

After incubation, excess antibody is washed away before adding the secondary antibody.

Question 10

What are common enzymes used in ELISA?

Answer 10

Common enzymes used in ELISA include alkaline phosphatase (AP) and horseradish peroxidase (HRP).

Question 11

What are the types of ELISA?

Answer 11

Types of ELISA include Indirect ELISA and Sandwich ELISA.

Question 12

What is the difference between Indirect ELISA and Sandwich ELISA?

Answer 12

Indirect ELISA uses a secondary antibody that carries the enzyme, while Sandwich ELISA uses capture and detection antibodies, making it more sensitive.

Question 13

What are some applications of ELISA?

Answer 13

Applications of ELISA include detecting cancer-related markers and infectious diseases.

Question 14

What can high levels of proteases in cancer lead to?

Answer 14

High levels of proteases can lead to degradation of fibrin, and these breakdown products can be detected by ELISA.

Question 15

What should be done after a positive ELISA result?

Answer 15

A positive ELISA result should lead to further tests to verify the findings.

Question 16

What is an example of a self-test kit?

Answer 16

PSA tests for prostate cancer.

PSA is an enzyme secreted by prostate epithelial cells.

Question 17

What do high levels of PSA in the blood indicate?

Answer 17

High levels (>4 ng/ml) may be a sign of prostate cancer.

When prostate cancer develops, the basement membrane is disrupted, leading to leaky blood vessels and increased PSA in circulation.

Question 18

How are PSA levels detected?

Answer 18

Detected readily from blood samples.

Question 19

What is the mechanism of the PSA test?

Answer 19

An antibody sandwich on a test strip gives a visual signal.

Question 20

What are the considerations for self-test kits?

Answer 20

Widely available, but require patient counselling and follow-up due to potential pitfalls of false positives.

Question 21

Can self-test kits be effective?

Answer 21

Yes, for example, COVID-19 self-test kits.

Question 22

What is Immunohistochemistry (IHC)?

Answer 22

Detects antigens in tissue samples.

Question 23

What components are used in Immunohistochemistry?

Answer 23

Uses primary and labelled secondary antibodies.

Question 24

How is visualization achieved in Immunohistochemistry?

Answer 24

Visualisation via HRP, AP, or fluorescent tags.

Question 25

What does a HER2 score of 0–1+ indicate?

Answer 25

HER2 negative.

Question 26

What does a HER2 score of 2+ indicate?

Answer 26

Borderline (FISH follow-up).

Question 27

What does a HER2 score of 3+ indicate?

Answer 27

HER2 positive → candidate for Herceptin.

Question 28

What is automated histological analysis?

Answer 28

It automatically scans clinical samples and ranks the extent of staining.

Question 29

What is flow cytometry?

Answer 29

A method for detecting specific molecules on and within cells.

Question 30

What can flow cytometry be used for?

Answer 30

It can sort/isolating specific sub-populations of cells.

Question 31

What is flow cytometry's role in cell biology research?

Answer 31

It is a powerful analytical technique.

Question 32

What does flow cytometry measure?

Answer 32

It measures surface/intracellular molecules on cells.

Question 33

How many antigens can be quantified simultaneously in flow cytometry?

Answer 33

Multiple antigens, up to 18 colours + FSC/SSC (cell size/complexity).

Question 34

What types of molecules are mainly used in flow cytometry?

Answer 34

Mainly proteins, but DNA and RNA can also be detected.

Question 35

What are some applications of flow cytometry?

Answer 35

Used for immunophenotyping, diagnosis (e.g. leukaemia), and basic research.

Question 36

What is cell sorting used for?

Answer 36

Used for research and therapy.

Question 37

What is High-Throughput Detection?

Answer 37

High-Throughput Detection involves techniques like Luminex Technology and Electrochemiluminescence for detecting multiple analytes in a sample.

Question 38

What is Luminex Technology?

Answer 38

Luminex Technology uses multiplex assays to detect a large number of proteins in a sample simultaneously.

Question 39

How does Luminex Technology capture proteins?

Answer 39

Fluorescently tagged beads capture specific proteins, with a secondary antibody having a PE (phycoerythrin) tag.

Question 40

How are beads read in Luminex Technology?

Answer 40

Beads are read in a flow-based system or, if magnetic, in a layer at the bottom of a well using lasers.

Question 41

What do the colours of the beads indicate in Luminex Technology?

Answer 41

The colour of the bead indicates the type of protein captured, while the colour of the PE indicates the quantity of the captured protein.

Question 42

What is Electrochemiluminescence?

Answer 42

Electrochemiluminescence is a method that offers high sensitivity and low volume for rapid quantification of multiple biomarkers.

Question 43

What is the CancerSEEK Blood Test?

Answer 43

CancerSEEK is a liquid biopsy test that detects dual biomarkers: mutations in ctDNA and protein biomarkers.

Question 44

What cancers can the CancerSEEK Blood Test detect?

Answer 44

CancerSEEK detects 8 major cancers with 60–100% accuracy, including stage I detection (varies by cancer type).

Question 45

What is the cost of the CancerSEEK Blood Test?

Answer 45

The cost of the CancerSEEK Blood Test is approximately $500.

Question 46

What are the limitations of the CancerSEEK Blood Test?

Answer 46

Limitations include being tested on known cancer patients and needing validation for early screening in general populations.

Question 47

What is the role of Liquid Biopsy in practice?

Answer 47

Liquid Biopsy allows for non-invasive detection of targetable mutations and guides therapy choices while avoiding repeated tissue biopsies.

Question 48

What is cfDNA and its significance in cancer detection?

Answer 48

cfDNA is normally present in serum and is generally unmethylated; however, methylation patterns change in tumours and can indicate cancer presence and tissue of origin.

Question 49

How accurate can cfDNA methylation patterns be in cancer detection?

Answer 49

Combined with machine learning, cfDNA methylation patterns can achieve accuracy up to 93%.

Question 50

What is an example of a test that uses cfDNA methylation patterns?

Answer 50

An example of a test that uses cfDNA methylation patterns is Galleri.