Serology

Station 1)
You are given the result sheet of chemilumences ( controls sheet or sample result, architec , vitrus )
Q1: How can you validate test results?
A1: by 1) checking the testing kit validity by expiry date - last opening date and condition of storage) all
according to manufacturer instructions.
Checking the test procedure by
A) check maintenance sheet to be sure status of device is working properly
B) Controls are within accepted range
e.g syphilis negative control : 0.035 and range ( -0.294 to 0. 294 ) so syphilis negative control is accepted
Syphilis positive control is : 2.706 and rang (1.916 - 3.944) so syphilis positive control is accepted.
Check the kit performance by QC sample results within the accepted range (LCL - UCL ) provided from
MRL.
After checking all previous steps and parameters are accepted the conclusion is RUN IS VALID
2) The check sample results:
If sample ratio (sample OD/cut-off) > 1 ===> initially reactive
Applying gray zone if sample ratio (HCV 0.8 - 1.0 } or (HBV , HIV, $ 0.9 - 1.0 ) result is INITIALLY REACTIVE
if sample < 0.8 in case of HCV ab and < 0.9 in case of HBsAg , HIV , syphilis Ab So ===> NON REACTIVE
initial serology screening
Non- reactive initially reactive
sample to be repeated in
DUPLICATE, SAME (sample-technique)
Result is Non reactive

Both NR -,- One R & One NR - , + both R +,+

Send sample for NAT
Result reported as Reactive
Discard blood unit and all its related components
You are give a NAT result sheet :

Q1: how to Validate the given results

A1: By A) checking the testing kit validity by Expiration date and storage condition
B) Checking the test procedure by Device validity
(maintenance sheet checked at proper time schedule
C) Check Calibrators (controls) ===> negative & positive controls should be valid
When all above criteria are accepted so ===> RUN IS VALID
D) Check the sample result : Note ( NAT cut off = 1, and sample ratio = (sample OD/cut-off)
Sample ratio > 1 =====> Reactive Sample ratio < 1 =====> Non- Reactive

Q2: How to interpret the obtained results?

A1: if initial serology screening test is NON_REACTIVE ==> perform NAT screening
when NAT result is non reactive ==> Release blood units and all related components for issuing.
* NAT is Initially Reactive is pooled NAT perform discriminatory NAT , the release result.
Other wise Report result as Positive for NAT ==> Discard blood units and all its related components
& permanent defer the donor from donation pool.

Questions on station 1 and 2

Q1: what are the types of NAT
A1: PCR & TMA , PCR (polymerase chain reaction) , TMA (transcription mediated amplification)
Q2:what are enzymes used in TMA NAT?
A2: RT( reverse transcriptase) , RNAP (RNA polymerase)
Q3: define ( NAT yield ) and (serology yield )
A3: NAT yield means tested sample is reactive with NAT while Non reactive with serology screening
 Serology yield means tested sample is reactive by serology screening but non-reactive by NAT.
Q4: what is window period and how NAT helps to minimize it?
A4: window period is the time from infection to test reactivity
NAT minimize it as follow : C, B, HIV, = 7, 8, 9.

Station 3
How to validate blood grouping results for donors?
By 1) checking the kits expiration date , date of last usage and storage condition
Including : ( blood group cards, AHG cards, anti-sera, and pooled screening cells)
3) checking machines and devices maintenance and performance according to SOPs and maintenance
sheet including , fully or semi-automated CAT devices (incubators - centrifuges) , sample tube centrifuge,
pipettes and refrigerators.
4) QC of the used kits and reagents
A) Blood grouping cards using A neg, B neg , AB neg , and O positive suspensions to verify that
cards can detect antigens properly.
B) AHG cards using negative and positive control serum against o positive cell suspension.
C) Pooled screening cells using positive and negative control serum
D) Anti-D anti-sera using (A neg, B neg , AB neg , and O positive suspensions) +Anti-D in AHG card

2nd Q :What is the flow chart of blood group sample in serology Lab?

MRL
Station 1) : using the provided Elisa kit for ( Hcv Ab , HBsAg , HIV AG/Ab, or syphilis) construt your
working sheet & monitoring sheet for the sample given:
 ⺁ ⺁ 폸኿⺁ ⺁ Ϯ Ϡ τ⺁ Β Θτ ΒΘ ⺁ Β Ϯ
Name of the assay
Reagent preparation ⺁ Β τ⺁ ኿Ϯ τ⺁ Ϯτ⺁ Ϯ ⰞΒ Ⱎ
Procedure ኿ΘΘ ⺁ Ro ‫ ف‬Β Β኿Ϯ ⺁ ϮΘ ኿ ⸱Β
Incubation time and temperature ⺁ ⺁ ϮΘ Ϯτ⺁ ⺁ R ⺁ ⺁ ⰞΒτ⺁
Stop , and reading wave length ⺁ τ⺁ Ϯτ⺁ Ϯ Ϡ τ⺁ ⸱

Work sheet : ⺁Θ Β኿ ⺁ R Β ⺁ ⺁ Β⸱ τ⺁ ⸱ τ⺁ ϠR Βτ
Monitoring sheet : ϮΒ ⺁ ⺁ R ⰞΒτ⺁ R ΒliΒ τ⺁ ⺁ ⺁ Β ⺁ ⺁ϮϠ Βτ ⺁
⺁R m ⺁ Β ኿ 10:00 Β 9:00 Β Θ툘Β ⺁ Ⱎ R Θ Β Β lΘ RϮΘ Ϯ ⰞΒ ⰞΒΒ ⺁
l Θ Ϡτ⺁ ⸱ ‫ف‬l ኿ �
Ϡ i τ⺁ ኿ ⸱䁕 Β ⺁ Ⱎ኿ R R ⺁ τ τ⺁ Β ኿ ⺁lR ⸱኿⺁ iΒτ⺁ ⺁ ⰞΒτ⺁ ⺁ ⸱τ Θ l ኿
Ϯτ⺁ Ϯτ⺁ Ϯ ⰞΒ Ⱎ ⺁ Θ툘Β
*Wash 20X means 1: 19 , wash : distilled water, if we need 200ml ===> 10 wash : 190 DW
If we need 400ml ===> 20 wash : 380 DW

* conjugate and substrate if needed preparation , spot method of preparation in pamphlet.

⸱ τ⺁ l Θ R ⺁ ⺁ Β⸱ ΒR Ϯ τ⺁ ⸱τ⺁ l Θ Β⺁ Ϯ Ϯ ⸱ ϮRϮΒ Ϯτ
Station 2): you are provided by print-out of an ELISA for test , validate the test and interpret results.
You will be given kit pamphlet and test kit
Use test pamphlet to have VALIDATION CRITERIA , and write it in details in your answer sheet

In the above part of pamphlet , I spotted validation criteria as follow:

* blank valid if between ( -0.020 , 0.050)
* NC should be < 0.120
* PC should be > 0.800
If validation criteria is met , so the RUN IS VALID ,
Proceed to calculations of Mean NC, the calculate the cut-off using the constant provided
Cut-off = Mean NC + 0.6 then calculate gray zone = cut-off X 0.8 because this HCV kit
Gray zone for HIV, HBsAg ,syphilis = cut-off X 0.9

** QC sample is placed following controls or as the last sample of the plate

QC ratio should be calculated (OD/cut-off) and then compared to reference range provided.

Station2)
You are given results for QC samples for Biorad HBsAg:
Day 1 2 3 4 5 6 7 8 9 10
Ratio 4.9 5.2 4.6 5.7 6.1 2.8 3.4 6.1 5.2 7.1
Provided that SD = 0.7
Draw a plot the graph.

Answer: Calculate Mean and SD (standard deviation) of the results

Mean = sum of all results divided by their count number= 51.1/10 =5.1
If one SD= 0.7 so 2SD = 1.4, and 3SD= 2.1
and upper control level = mean+ 3SD = 5.1+2.1= 7.2 & LCL= mean - 3SD=5.1-2.1=3.0
⸱τ⺁ lⰞ ኿ τ⺁ ኿τ⺁ Β
⺁ R኿ Β Ϯ τ Θ ϠΒ 䁕 Β Θτ⺁ Ϯ m 7.2 3.0 ⺁ Θ R Β⸱τ⺁ Β l ⺁ Ϯτ
Scenarios to say that I have invalid QC results
1) Outside + or - 3SD 2) 5 or 7 points on one 3) Trend 4) Cyclic
side of the Mean ኿ ⺁ τ ϮΒ τ⺁ τ⺁Ϯ ኿ τ
Causes of invalid QC results
Sample related causes Procedure related causes
1) Improperly stored/ transported 1) expired kit
2) Contaminated 2) Non- maintained / un-calibrated devices
3) Preservation temp. Is not maintained 3) Missed steps in procedure
4) Deteriorated (repeated thaw/ freeze cycles) 4) Non trained operator
5) Expired sample 5) Carry over by washer
6) Failure to interpret with proper kit

You are given results for QC samples for Biorad HBsAg:

Day 1 2 3 4 5 6 7 8 9 10
Ratio 4.9 5.2 4.6 5.7 6.2 2.8 3.4 6.1 5.2 7.1
Provided that SD = 0.7
Calculate mean , mode , median
Mean = sum of values/number of values= 51.1/10= 5.11
Mode = most frequently repeated value = 5.2
Median = middle value between ascending or descending arrangement (if odd number of values)
= mean of 2 middle values betweenascending or descending arrangement (when even number)
7.1 , 6.2 ,6.1 ,5.7 ,5.2 ,5.2 ,4.9 ,4.6 ,3.4, 2.3
5.2 2 5.2 5.2 Ϡ ϮΒΒ ⸱m τ⺁ Β iτ⺁ τ⺁ m ⸱ R ኿ ኿l⸱
So median = 5.2

Infection control
Q1:what are the moments in your practise that require hand hygiene ?

Q2: what are missed parts of hand during hand wash?

Q3: face mask is one of PPEs that require special care during putting on and taking off
Mention do and don’t with face mask

DO DON’T

Wear a mask that covers your nose and mouth
* Wash your hands before putting a mask on
*Put the mask on using the straps instead of
touching the cloth
*Remove the mask by the straps
* Wash your hands after you take it off
Store it in a clean, dry container
Wash your mask daily in hot water
Hold onto the cover when you apply your mask
Adjust or touch the mask once it’s on properly
Slide it down under your chin
Wear it if it’s soiled, wet, ripped, or damaged in
any way
Share your face mask with someone else

Q3: waste disposal and management steps

1) Segregation At point of origin in special colored bag
2) collection By janitors wearing duty gloves and foot wear
3) storage Dedicated place -impermeable- good drainage - cleanable surface
4) transportation Dedicated vehicle out of facility
5) Treatment Either by incineration or chopping and sterilization
6) Final disposal Burring in sanitary landfill (solids) or pouring in special sinks (liquids)

Q4: during your work in components lab department , blood bag ruptured and caused blood spill
How janitors should respond to large blood spill and small blood spill
1) Wear full protective PPE Small spill
2) Remove blood with towel , cotton , or tissue 1) Wear protective gloves
paper, the discard it to red waste bag. 2) Sweep the surface several times with
3) Pour disinfectant (chlorine 5000 ppm on site of disinfectant and tissue paper
spill and leave for 10 minutes. 3) Finally clean the surface.
4) Dry disinfectant with tissue paper
5) Finally clean the surface

Q6: how to prevent and minimize the risk of blood born infection in your lab?
A6:
1) Promote HBV vaccination among health care workers.
2) Apply the standard precaution of infection control.
3) Prevent per-cutenous injuries by implementing safe injection policies and procedures.
4) Avoid needle recapping
5) Apply the work practise ethics.
6) Using the engineering controls.

Q7: one of your team is exposed to blood , how to manage ?

A7:
1) Wound care CLEAN BY SOAP & WATER NO milking , no antiseptic no , disinfectant at wound
2) Fill an exposure report.
3) Determine risk associated with the exposure (type of fluid- type of exposure.)
4) Evaluate exposure source.
5) Give PPEs to personnel involved in risk of exposure to infection transmission.
6) Provide counselling for the exposed person.
7) Perform followup testing.

Q8: what are levels of risk of blood borne infections?

HIV ------> 0.2 - 0.4 %
HCV ------> 3 - 10 %
HBV ------> 27 - 37 %

Q9: COVID 19
폸⸱τ⺁ Β ⺁ R ⰞR τ⺁

RCRL
Station 1) Ab screening and id identification
Β኿Ϯ Ϯ ⺁ ϮΒ Β ኿ ⺁ � 11 Β 1 Β R Β R ml Θ � R τ ኿ R R Ϡ Βm
τ ⺁ ΘΘ ab screening
 0 ⺁4 Β⺁ ⸱R Θ Θ Βτ⺁ Ⱎ ⺁ τ⺁ Ϯ Βm
⺁ τ⺁ R Β⸱τ⺁ Β⺁ Ϡ኿Βτ⺁ ‫ف‬l ኿ Rl Θ ⺁ R Β኿ ⺁ ⺁ R Β⸱τ⺁ ኿⺁
Β⸱τ⺁ l኿ τ⺁ Ϯ኿኿ Rl Θ

Follow up questions will be like those:

1) What is the most probable antibody the patient has?
2) What antibodies can`t be excluded? What further steps should be taken to exclude them?
3) If patient needs blood , what unit phenotype should be provided?
4) An embedded question to test RH phenotype nomenclature
Station 2: ABO discrepancy case = the same gel card and you will observe grads of reaction
Questions will be like this:
1- What is most probable cause of this discrepancy ?
2- What are the causes of Abo discrepancies?
3- What is the most common cause of group discrepancies:
4- How to resolve a case of ABO discrepancy?
Q1: how to validate anti-sera?
A1: by checking its
specificity Sensitivity stability Avidity ( speed of reaction) Potency (by titration)

Q2: How to validate a blood group card?

A2:
1) By checking its reactivity in various blood groups using
(Aneg , B neg , AB neg and o psitive cell suspensions)
2) Checking its reactivity in subgroup reactions e.g A2
3) Checking its reactivity in case of weak D