Early Infant Diagnosis

using DBS-PCR

Collection, Drying, Package,

and Transport of DBS
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 What Is a Dried Blood Spot
(DBS)?
• Whole blood dried on filter
paper
• Usually obtained from pricking
skin, not from phlebotomy

2
What Is a Dried Blood Spot
(DBS)?

3
 What Is a Dried Blood Spot
(DBS)?
• Requires only a small amount of
blood
– 50ul - 80ul is needed per spot
• Easy to store
– Stable at room temperature for more
than 90 days
• Easy to transport
– Envelope
• Reduced biohazard*
4
Why to use DBS in EID Programs?
• DBS testing requires smaller volumes of
blood (avoids phlebotomy)
• Are less infectious and not prone to glass
breakage, and have no need for cold
storage during transportation.
• DBS testing for infant diagnosis can be
done with commercially available DNA
PCR with high sensitivity and specificity
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 Why to use DBS in EID Programs
cont………
• Providers can be trained to collect these
samples and properly store them for
transport (simplifies specimen collection
process).

• DBS dramatically simplified access to HIV testing, it was

adopted by the national HIV/AIDS program as means of
specimen collection for infants and young children .

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 .

How to collect DBS

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 What Are The Responsibilities of
The Trainee?
• Label specimens correctly
• Fill in lab request forms and records
correctly
• Collect valid specimens *
• Store specimen appropriately until
transported for testing
• Facilitate referral of specimen
A test result is only as good as the
specimen collected
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 Required Supplies for DBS
collection*
• Powder free gloves
• Pen
• Lab request forms &
HEI register
• DBS card
• Lancet
• Disinfectant for skin
• Gauze or cotton wool
• Waste container

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 How to Collect DBS
• Use Universal Safety
Precautions for
handling blood*
• Clearly label card with
appropriate
identification number
before collecting the
blood.
• Do not touch circles
with your fingers, or
anything else**
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 Steps of Toe/Heel prick DBS
Specimen Collection 2nd
choice*
1. Mother warms the foot
2. Wash hands, put on gloves
3. Position baby with foot down
4. Clean area, dry 30 sec
5. Press lancet into foot, prick
skin
6. Wipe away first drop
7. Allow large drop to collect
8. Touch blood drop to card
9. Fill entire circle with drop
10. Fill at least 4 circles
11. Clean foot, no bandage 1st choice*

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12
Warm the area*

13
Wash hands and put on glove

14
Position baby with foot down*

15
 .

How to collect DBS through toe

stick

16
Clean the area, dry 30 seconds*

17
Press lancet into foot, prick skin*

18
Wipe away first drop*

19
Allow large drop to collect

20
Fill entire circle with drop*

21
Fill at least 4 circles

22
* Collecting more than 4 spots is
preferred

23
Clean area, do not use bandage*

24
 Technique of holding and
squeezing the foot during toe stick*

Correct way Incorrect way 25

 .

How to collect DBS through

heel stick

26
Warm the area

27
Position baby with foot down

28
Clean area, dry 30 seconds

29
Press lancet into foot, prick skin

30
Wipe away first drop

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Allow large drop to collect

32
Technique of holding & squeezing
the foot during heel stick*

33
Touch drop to card

34
Fill at least 4 circles

35
Clean foot, leave with no bandage

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 Steps of Toe/Heel prick DBS
Specimen Collection 2nd choice*
1. Mother warms the foot
2. Wash hands, put on gloves
3. Position baby with foot down
4. Clean area, dry 30 sec
5. Press lancet into foot, prick
skin
6. Wipe away first drop
7. Allow large drop to collect
8. Touch blood drop to card
9. Fill entire circle with drop
10. Fill at least 3 circles
11. Clean foot, no bandage
1st choice*
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How to Inspect the
quality of DBS at site
level

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What is a valid DBS specimen?
• Proper identification
– date of collection, name, date of birth, unique
identifier (region/type of facility /facility ID/
unique ID)
• Number of circles (4 or more spots)
• Completely filled spots
• Completely dried spots
• No sign of hemolysis, clotting, scratching or
contamination
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Valid DBS specimen

40
Not valid. Note blood drops too
small with toe squeeze

41
Invalid DBS Specimens
Packed before
drying

Hemolysis

Clotted/ layered

Incomplete circles
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 .

DBS Quiz

43
Is this DBS #1 acceptable?

44
 Answer to Q1

Preferred DBS card with 5 properly

collected samples

-Properly collected DBS samples completely filling the

circles
-Clearly readable identification information

45
Is this DBS#2 acceptable?

46
 Answer to Q2

Acceptable DBS card showing 4

well collected samples

Guideline states that a minimum of 4 spots be

-

prepared that completely fill circles

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Is this DBS #3 acceptable?

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 Answer to Q3

• Not preferred DBS card with 5 incompletely

filled circles
• Blood must fill up the entire circles, to
provide a 12 mm punch needed for testing.

49
Is this DBS#4 acceptable?

50
 Answer for Q4

Rejected DBS card due to inadequate ID

information

-Should not reject it outright because DBS is acceptable.

- Contact the collection site manager to obtain missing
information.

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Is this DBS#5 acceptable?

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 Answer to Q5

Rejected DBS card, showing halo around

the DBS due to the contamination of
alcohol
-Clean the site for sample collection with alcohol
swab and allow it to DRY thoroughly before .
-This DBS is NOT suitable for PCR test because
sample is diluted with alcohol.

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Is this DBS #6 acceptable?

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 Answer to Q6

• Rejected DBS card showing inadequate

amount of blood collected within the
circles.
• Pricked skin was touched immediately on
to a DBS card without waiting for full drop
to form.
• Note: A full drop of blood must form before
touching the drop to the card.
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How to dry, store, and
transport DBS for the
testing PCR-LAB

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 How to Dry DBS
• Don’t touch or smear the blood spots
• Allow the specimen to air dry horizontally
for at least 3 hours.
• However, you keep the DBS overnight in
case there is high humidity in the room.

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 How to Dry DBS*….
• Keep away from direct sunlight, dust, and
bugs
• Do not heat, stack or allow DBS to touch
anything during the drying process

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Dry Completely Before
Packaging

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 Required Supplies to Package DBS

• Proper Dried DBS

• Weighting paper
• Sealable
impermeable plastic
bags
• Desiccant packs

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How to Package DBS for Storage
1. At the end of each day, pack the dry DBS (leave for
next day if not dry 3 hrs)
2. Wrap DBS card in a weighting paper.
3. Insert into sealable plastic bag
4. Add 1 desiccant per DBS card in the bags
5. Add 1 humidity card per bag
6. Press air out of bag
7. Label bag with facility code, range of unique
identifiers, and seal
8. Put in an envelope at room temperature in darker
place if it is not going to be shipped to the lab that
day
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Insert Cards into Sealable Plastic
Bag Warped with Weighting paper

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3. Add Desiccant Packets
Minimum 1 per DBS specimen per bag*

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4. Add Humidity Cards and Seal
Bag

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 How to Store DBS
Keep packaged DBS (in sealable plastic bags)
at room temperature until transported to
reference laboratory
It does not need of refrigeration

Avoid leaving in
vehicle, as sun and
heat will deteriorate
DBS

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 How to Package DBS for
Shipping
1. Inspect the humidity
indicator (if the humidity is
> 50%, replace the
desiccant packets)
2. Insert into envelope
3. Include lab requisitions
4. Label outside clearly
5. Date of packing
6. Send to PCR lab

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 Send Soon to regional
laboratory..!!!!
Thank you for participating!

Any last questions?

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