Brainscape's Knowledge GenomeTM

Browse over 1 million classes created by top students, professors, publishers, and experts.


See full index

3. Transfusion > IBTS - Nucleic Acid Testing > Flashcards

IBTS - Nucleic Acid Testing Flashcards

1
Q

How many people receive BTs every week?

A

Over 1000 every week

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

How many donations do we need every week to fulfil requirements?

A

3,000

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

How many donations so we get every year?

A

140,000

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

Write about using serology and NAT testing to test our donors

A

NAT and serology complement each other to detect viral infections

We use Alinity analysers in serology but Panther analysers in NAT

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

What do we test for in NAT?

A

HIV-1/2
HCV
HBV
HEV
Selective WNV testing

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

Is HIV RNA or DNA

A

RNA

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

Is HBV RNA or DNA

A

DNA

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

Is HCV RNA or DNA

A

RNA

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

Is HEV RNA or DNA

A

RNA

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

Is WNV RNA or DNA

A

RNA

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

Why use NAT

A

NAT detects very low levels of viral RNA or DNA
They are qualitative tests -> either positive or negative (not great for donor as it doesn’t say how much of an infection they have)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

What is meant by selective West Nile Virus testing?

A

Only test on donors who have travelled to a risk area

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

What is the newest assay introduced

A

Hepatitis E assay

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

When can NAT not detect a virus?

A

Cannot detect viral infections during the eclipse phase (when stored in liver/lymph)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

What sample type is used for NAT

A

Plasma or serum

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

What is the window period

A

The time between acquiring an infection and when it can be detected using serological markers (antibodies)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

What is the main benefit or NAT?

A

Detects viral RNA or DNA early in the development of infection (during the window period) before antibodies are detectable

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

Comment on RNA replication versus DNA replication

A

RNA replicates rapidly -> substantial increase in viral load <5 days after infection -> doubling time only a few hours e.g. HIV @ 20 hours

DNA replicates slower -> can take several weeks to reach detectable levels e.g. HBV which has a doubling time of 2-3 days

19
Q

Write about the combination of HBV NAT testing and HBV serology testing

A

HBV DNA can detect early infection (3 to 12 weeks)
HBV DNA + HBsAg detects acute infection (3 to 12 weeks)
HBV DNA + anti-Hbc total - detects chronic infection (DNA might not be detectable)

20
Q

What proves to be the biggest risk to donors and why

A

HBV due to its very slow doubling time making it very difficult to detect

Low DNA levels are undetectable during the window period or are often below the limit of detection by the assay

Low DNA also seen during chronic HBV infections which can also be HBsAg negative -> may only have anti-Hbc positive serology

21
Q

What is a chronic HBV infection also called?

A

Occult HBV infection

22
Q

What Assay method is used

A

Ultrio Elite Assay Methodology

23
Q

What are the main steps to the Ultrio Elite Assay method?

A
  1. Sample prep -> DNA/RNA extraction
  2. RNA/DNA target amplification
  3. Detection of amplified target
24
Q

How is DNA/RNA amplified using the Panther?

A

Transcription mediated amplification (TMA)

25
Q

How do we detect the amplified products using the panther assay?

A

Using the hybridisation protection assay (HPA) and dual kinetic analysis

26
Q

Describe how the assay works

A

The target capture reagent (TCR) contains a detergent which solubilises the viral envelope - DNA/RNA now available

Oligonucleotides (primers) are added which bind to the DNA/RNA

The oligo/NA complex binds to magnetic particles in the TCR which allows for the isolation of the NA

A TER (target enhancer reagent) can be added which enhances the disruption of HBV viral particles

27
Q

How does transcription mediated amplification work

A

An enzyme reagent is added containing two enzymes:
- MMLV reverse transcriptase
- T7 RNA polymerase

28
Q

How does MMLV Reverse transcriptase work?

A

Generates DNA copy of the target sequence

Containing a promoter sequence for T7 RNA poly

29
Q

How does T7 RNA polymerase work?

A

Produces multiple copies of RNA amplicon from the DNA copy template

30
Q

How does the detection step of the assay work

A

A probe reagent is added containing a chemiluminescent label which binds to hybridised probe + virus to give a positive result

A selection reagents is added which inactivated any unhybridised probes (probes not bound to viral NA)

The chemiluminescent signal from the probe is measured in a luminometer and reported as relative light units (RLUs)

31
Q

How does dual kinetic analysis work?

A

There is 2 results -> an internal control or a ‘flasher’ signal and the sample ‘glower’ signal -> both are required for the test to be valid

Results are graded as reactive, non reactive (negative/no virus), invalid (control failed - no ‘flasher’)

32
Q

How do we test for multiple viruses at once

A

Multiples UE assay
Tests for all three viruses at once

33
Q

What do we do if we get an initial reactive (IR)/positive result?

A

Our first test is a multiplex UE assay -> tests for 3 viruses at once

If positive we need to run a disciminatory assay

34
Q

What is a discriminatory assay?

A

An assay whereby we use discriminatory probes to determine which virus is present i.e. which are repeat reactive in the sample

Tells us if its HIV, HCV or HBV

35
Q

Why do we not have disciminatory probes for HEV and WNV?

A

These two assays only test for HEV or WNV, => no need for discrimination

36
Q

What do we do if all three disciminatory probe assays come up positive?

A

Need to retest the donation -> very low chance that donor is positive for all three at once -> if still positive probably reacting against some component of the assay

37
Q

What is West Nile Virus

A

A mosquito borne flavivirus transmitted in wild and captive birds

A neuropathogen which affects humans, equine and avians

Single stranded RNA virus lineage 1 and 2

38
Q

When do we decide to test for WNV

A

We check the ECDC map every week to determine which countries are considered high risk

Anyone coming from these countries needs to be tested

39
Q

In general when do we test for WNV

A

Seasonal testing done between may and december on those from high risk areas

We have not had a positive test yet

40
Q

Why do we test for WNV even if we havent seen a single positive yet?

A

We can accept additional blood products collected from donors who otherwise would have been deferred
- think about rare phenotypes in SCD that could need this blood

41
Q

Talk about HEV testing

A

The HEV assay is the only assay directly funded by the department of health - brought in from 2016

Incidence of about 1 in 5,000 Irish donations

1-4 genotypes

Transmitted through domestic pigs, consumption of undercooked pork and shellfish

42
Q

Why is HEV testing so important in Ireland?

A

in 2020 there was 58 repeat positives -> meaning there was a 1 in 2,314 chance of HEV

There is no consistent seasonal pattern of HEV infection so all donors are tested

43
Q

What sample types other than plasma/serum are seen in NAT lab?

A

Tissue samples
Maternal/Cord samples
Bone marrow samples
Virology positive follow ups

44
Q
A

3. Transfusion (14 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2024 Bold Learning Solutions. Terms and Conditions