BLOOD COLLECTION METHODS AND PROCEDURE

1. BLOOD COLLECTION
NEETHU SOMAN
MSc Medical Biochemistry

2. COLLECTION OF BLOOD
• Blood collection is three types.
1. Arterial blood collection : Carrying oxygenated blood from heart to several
parts of the body by the pumping action Of heart.
2. Venous blood collection: Carrying deoxygenated blood from distant parts of
the body to heart & to lungs.
3. Capillary blood collection: a minute blood vessel that connects the smallest
arteries to the smallest veins and serves as an oxygen

3. CAPILLARY BLOOD COLLECTION
• Capillary puncture, also called dermal puncture, is a safe, rapid, and efficient
means of collecting a blood specimen.
• To perform capillary puncture, a small sterile lancet or blade is used to puncture
the skin and capillaries to create a blood flow.
• Capillary punctures are performed when only a small amount of blood is required,
when obtaining blood from infants, or when the patient has a condition that makes
venipuncture difficult.
• In the clinical laboratory, capillary blood has been used only in special situations,
because of the small sample volume and potential for rapid clotting of the sample.
• However, the increased use of small, portable, easy-to-use instruments that
require only a drop or two of blood, has made capillary blood the specimen of
choice for these analyzers.

4. Capillary Puncture Sites
• The usual site for capillary
puncture in adults and children is
the fingertip .
• In adults, the ring finger is often
selected because it usually is less
calloused.
• For newborns and infants, capillary
blood can be obtained from the
lateral or side portion of the heel
pad. Once an infant begins to walk
(about the age of 1 year) blood
should be collected from a
fingerstick.

5. Capillary Puncture Equipment
Lancets : Several designs of disposable,
sterile safety lancets are available for
capillary puncture.
• These lancets make punctures of
uniform depth at the touch of a button
and are available in several blade
lengths for use in different situations.
• Special pediatric lancets that produce a
shallow puncture should be used with
infants.

6. Capillary Blood Collection Equipment
• The following are needed for routine capillary blood collection:
1. Sterile lancet—Retractable type is preferred; tip less than 2.4 mm. The tip length will depend
on the intended use. For newborns and infants under 12 months, the device should cut or
puncture no more than 0.85 mm. The lancet be inserted at during capillary blood collection at
45 degrees .
2. Alcohol wipes—Wipes containing 70% isopropyl alcohol.
3. Sterile gauze pads—For removal of first free-flowing drop of blood and for pressure
application after collection.
4. Gloves—Can be made of latex, rubber, vinyl, etc.; worn to protect the patient and the
phlebotomist.
5. Eye protection—To protect the phlebotomist from aerosols contaminating the eyes.
6. Sharps disposal unit—Lancets must be placed in a proper disposal unit immediately after
their use.

7. • Capillary Collection Containers : Capillary
blood can be collected in capillary tubes or
collection vials. Capillary blood required for
tests other than microhematocrit, such as
chemistry tests, can be collected in special
vials with a capillary or other extension for
directing the blood into the vial .
• These vials are available plain or with
anticoagulant. Using these vials, a small
quantity of whole blood, plasma, or serum
can be obtained

8. Collecting the Blood Sample
• The first drop of blood should be wiped away with dry, sterile gauze.
• This first drop contains tissue fluid, which dilutes the blood drop and can also activate
clotting.
• The second and following drops of blood are used for the test sample.
• A well-rounded drop of blood should be allowed to form before collection begins.
• The hand can be gently massaged to increase blood flow, but excessive pressure near the
puncture site should be avoided. (Squeezing the fingertip can force tissue fluid into the blood
sample.)
• Capillary blood should be collected as quickly as possible to prevent clotting.
• The capillary tube should be held in an almost horizontal position, or tilted slightly downward;
the blood collecting vial should be held vertically so blood will flow down into the tube.
• When the tip of the capillary tube is touched to the blood drop, blood will enter the tube by
capillary action because of the attraction between the liquid and the tube.
• Capillary tubes should be filled three-quarters full

9. PERFORMING THE CAPILLARY PUNCTURE
(A) perform puncture on
cleansed fingertip; wipe away
first blood drop and
(B) allow rounded drop of
blood to form;
(C) collect blood into
capillary tube or
(D) collection vial

10. Caring for the Capillary Puncture Site
• After the blood has been collected, sterile gauze or a cotton ball should
be placed on the puncture site and pressure applied until bleeding
stops.
• A small adhesive bandage can be applied if necessary.

11. PRECAUTIONS
1. Always wear lab coat and gloves while collecting blood sample.
2. Do not squeeze puncture site too tightly as this can dilute the sample.
3. Avoid touching blood specimen and puncture site.
4. Puncture depth should be less than 1.5mm.
5. Use middle or ring finger of hand for capillary blood collection.
6. Never touch tip of the lancet.
7. Always use disposable gloves and disposable lancet.

12. VENOUS BLOOD COLLECTION
• Venipuncture is a common method of obtaining blood for laboratory
examination.
• The venipuncture is a quick way to obtain a large sample of blood on which many
different analyses can be performed.
• In a venipuncture, also called a phlebotomy, a superficial vein is punctured with
a hypodermic needle and blood is collected into a vacuum tube or syringe.
• The venipuncture is a safe procedure when performed correctly by trained
personnel.
• These personnel can include phlebotomists, medical assistants, nursing staff,
physicians, medical laboratory technicians, and medical laboratory scientists.
• The venipuncture must be performed carefully to preserve the condition of the
vein. Much observation and practice under the supervision of an experienced
phlebotomist is required to become skilled and self-confident in the art of
venipuncture

13. • Performing a venipuncture involves several important steps that must be
thoroughly understood before the procedure is attempted: Observing Standard
Precautions and other safety measures throughout procedure.
• Selecting the proper equipment
• Identifying the patient using two identifiers
• Preparing the patient for venipuncture
• Selecting and preparing the puncture site
• Applying and removing the tourniquet
• Obtaining the blood
• Caring for the puncture site
• Observing the patient for adverse reaction
• Labeling blood specimens immediately following blood collection

14. VENIPUNCTURE MATERIALS AND SUPPLIES
• Venipuncture can be performed using a safety needle/collection tube holder
assembly , a safety needle and syringe, or a winged collection set with tubing
and tube holder .
• Other materials required for venipuncture include evacuated blood collecting
tubes, alcohol swabs, sterile gauze,disposable tourniquet, and small adhesive
bandage.
• Venipuncture is performed using a vacuum-tube system, a syringe, or a winged
collection set. The safety guidelines and quality assessment considerations are
the same for all venipuncture methods.

15. VENIPUNCTURE MATERIALS AND SUPPLIES
1. Safety Needles, 22g or less
2. Butterfly needles. 21g or less
3. Syringes
4. Vacutainer tube holder
5. Transfer Device
6. Blood Collection Tubes.
• The vacuum tubes are designed to draw
a predetermined volume of blood.
• Tubes with different additives are used
for collecting blood specimens for
specific types of tests.
• The color of cap is used to identify these
additives.
7. Tourniquets. Single use, disposable,
latex-free tourniquets
8. Antiseptic. Individually packaged
70% isopropyl alcohol wipes.
9. 22 Gauze
10. Sharps Disposal Container. An
OSHA acceptable, puncture proof
container marked “Biohazardous”.
11. Bandages or tap

16. VENIPUNCTURE MATERIALS AND SUPPLIES

17. PROCEDURE
1. Identify the patient, two forms of active identification are required.
• Ask the patient to state their name and date of birth.
• This information must match the requisition.
2. Reassure the patient that the minimum amount of blood required for testing will
be drawn.
3. Verify that any diet or time restrictions have been met.
4. Order of Draw
• The following order of draw is the approved order as established by CLSI.
• This order of draw should be followed whenever multiple tubes are drawn during
a single venipuncture.
• This is to prevent cross contamination by the tube additives that could lead to
erroneous results

18. 1. Assemble the necessary equipment appropriate to the
patient’s physical characteristics.
2. Wash hands and put on gloves.
3. Position the patient with the arm extended to form a
straight-line form shoulder to wrist.
4. Do not attempt a venipuncture more than twice. Notify
your supervisor or patient’s physician if unsuccessful.

19. 5. Select the appropriate vein for venipuncture.
• The larger median cubital, basilic and cephalic veins are most frequently used, but
other may be necessary and will become more prominent if the patient closes his fist
tightly.
• At no time may phlebotomists perform venipuncture on an artery.
• It is not recommended that blood be drawn from the feet .The Providers permission is
required to draw from this site.
• Extensive scarring or healed burn areas should be avoided
• Specimens should not be obtained from the arm on the same side as a mastectomy.
• Avoid areas of hematoma.
• If an IV is in place, samples may be obtained below but NEVER above the IV site.
• Do not obtain specimens from an arm having a cannula, fistula, or vascular graft.
• Allow 10-15 minutes after a transfusion is completed before obtaining a blood sample

21. 6. Apply the tourniquet 3-4 inches above the collection site.
• Never leave the tourniquet on for over 1 minute.
• If a tourniquet is used for preliminary vein selection, release it and reapply after
two minutes.
7. Clean the puncture site by making a smooth circular pass over the site with the
70% alcohol pad, moving in an outward spiral from the zone of penetration.
• Allow the skin to dry before proceeding.
• Do not touch the puncture site after cleaning.

22. 8. Perform the venipuncture
A. Attach the appropriate needle to the hub by removing the plastic cap over the small end of the needle and
inserting into the hub, twisting it tight.
B. Remove plastic cap over needle and hold bevel up.
C. Pull the skin tight with your thumb or index finger just below the puncture site.
D. Holding the needle in line with the vein, use a quick, small thrust to penetrate the skin and enter the vein in one
smooth motion.
E. Holding the hub securely, insert the first vacutainer tube following proper order of draw into the large end of the
hub penetrating the stopper. Blood should flow into the evacuated tube.
F. After blood starts to flow, release the tourniquet and ask the patient to open his or her hand.
G. When blood flow stops, remove the tube by holding the hub securely and pulling the tube off the needle.
H. Gently invert each tube
Light blue top- invert 3-4 times
Red and gold tops invert 5 times.
All other tubes containing an additive should be gently inverted 8-10 times.

23. 9. Place a gauze pad over the puncture site and remove the needle.
10. Activate the safety device and properly dispose of the vacutainer holder with needle attached
into a sharps container.
11. Immediately apply slight pressure to the gauze pad over the venipuncture site..
• Ask the patient to apply pressure for at least 2 minutes.
• When bleeding stops, apply a fresh bandage, gauze or tape.
12. Tubes must be positively identified after filling with a firmly attached patient label.
• The label must include the patient first and last names, DOB,, collection date and time and
collectors initials.
• If no patient labels are available, manually label the tubes with the required information. All
labels must include two identifiers .
• The tube must be labeled before leaving the patient.
13. Observe special handling requirements
• Some test specimens require special handling for accurate results.

24. Anticoagulants and Additives
• The standard operating procedure (SOP) manual will include a list of tests performed and the
type and size of vacuum tube that should be used when collecting blood for each test. The cap
colors of the blood collection tubes designate which, if any, anticoagulant or additive is present
. When multiple tubes must be filled during a venipuncture, a particular order of draw, such as
shown , must be used.
• Filling tubes in the correct order prevents cross contamination of anticoagulants between
tubes, which could adversely affect test results. When the tube stopper or cap is pierced by the
blunt end of the venipuncture needle, the vacuum draws the blood into the tube. It is important
that tubes be filled to their stated capacities because an incorrect blood: anticoagulant ratio can
alter cell morphology and cause erroneous test results, especially in coagulation tests.
• Some tubes have a fill line marked on the tube to easily determine if blood volume is
sufficient. For certain tests, tubes with anticoagulant can still be used for tests if they are not
completely filled but are at least 70% full.
• The laboratory’s SOP manual will specify the tests for which the level of tube fill is critical.
For example, one requirement for coagulation specimen acceptability is that citrate tubes must
be filled to at least 90% capacity.

26. When collecting multiple specimens, blood tubes should be drawn in the
following order; mix all tubes by inversion 6 – 8 times:
1. Blood Culture bottles
2. Isolator tube
3. Blue top (3.2% sodium citrate)
4. Red top (no preservative) and Gold top (SST)
5. Royal blue top (no preservative)
6. Green top (sodium heparin)
7. Lavender top (EDTA), Pink top (EDTA), and Royal
blue top (EDTA)
8. Gray top (sodium fluoride)
9. Yellow top (ACD) Solution A or B
10. TB Gold QuantiFERON: Nil (gray top), TB antigen
(red top), and Mitogen (purple top)

27. Arterial blood Collection
• Arterial blood is the preferred specimen for Blood Gas determinations.
• Arterial punctures are technically difficult with potentially more risk for the
patient than venipuncture; therefore, in clinical practice, it is reserved for the
most essential purposes.
• Conditions that commonly utilize the measurement of blood gases include
chronic obstructive pulmonary disease (COPD), cardiac and respiratory failures,
severe shock, lung cancer, diabetic coma, coronary bypass, open hear surgery,
and respiratory distress syndrome (RDS) in premature infants.
• Patients requiring blood gas determinations are often critically ill and require
vigorous measures to support life through assisted ventilation using mixtures of
gases tailored, in response to laboratory findings, to serve individual needs.

28. SPECIMEN REQUIREMENTS:
• A standard blood gas collection kit is used in all areas. The collection
kit utilizes dry lithium heparin and a syringe volume preset collection
system.
• Routine practice is to preset the volume at 3 mL. for arterial blood gas
collections. and to expel any air in the syringe after sample collection
and to transport on ice to the central laboratory.

29. PROCEDURE:
Patient Preparation
A. The phlebotomist must wash their hands, prior to attending to the patient, with
either soap and water or “waterless soap.”
B. Patient identification is made by cross-checking patient orders against patient
arm band and room number for accuracy.
C. Patient is informed that the collection is an arterial puncture requested by their
physician. Inform them that the puncture could be more painful than a
venipuncture and that the site must be held with pressure for a at least 5 minutes.
If the patient is on anticoagulants, it may require additional holding time until
bleeding has completely stopped. If bleeding persist longer than 10 minutes
contact the patients nurse.
D. Obtain and record preliminary patient information

30. Site Selection
• A. Radial artery . The radial artery is the preferred site for arterial puncture as the
ulnar artery provides collateral circulation to the wrist and it is close to the surface
of the wrist and easily accessible. If both radial arteries are unacceptable for use
the next site would be the brachial artery.
• B. Brachial artery. This site is reserved for secondary use as the median and
ulnar nerves and the basilic vein are in close proximity to the brachial artery. The
femoral is the last site of choice
• C. Femoral artery. Infection is a risk at this site and it may be more difficult to
control the bleeding at this site post puncture. Pressure must be applied at this site
for a minimum of 10 minutes. Femoral punctures are only done by request of the
physician. They are performed most frequently during “Code 99” because it is
usually the easiest site to palpate and puncture .

31. Performing the Puncture
A. Prepare the syringe by setting the preset volume and positioning the needle with
the bevel side up.
B. Enter the artery at a 45 degree angle (radial) (90degree for brachial) slowly, until
blood appears in the needle hub. If blood does not appear, the needle may be
slightly redirected but remain under the skin. NOTE: If you are unsuccessful after
two attempts then withdraw the needle and allow the patient to rest. Allow someone
else to perform the puncture. Use a new syringe and needle with each attempt.
C. When the preset volume has been collected, remove the needle and apply firm
pressure for a minimum of 5 minutes (5 for radial or brachial, 10 for femoral
puncture) by placing the sterile gauze pad over the puncture.
D. Expel any air from the syringe, replace the needle with the syringe cap provided,
and rotate the syringe to insure thorough mixing of the sample with the dried
heparin.

32. LIMITATIONS OF PROCEDURE:
• SOURCES OF SAMPLE ERROR:
A. Air in the sample syringe -- The atmospheric oxygen enters the specimen and
carbon dioxide from the specimen enters the air bubble.
B. Clotted Sample -- Caused by inadequate mixing of the specimen with
anticoagulant after collection.
C. Delay in testing a sample not on ice -- Blood cell metabolism continues
utilizing oxygen and produces carbon dioxide and acids that lower pH.
D. Insufficient patient information documentation -- No patient temperature. No
oxygen therapy information. Site of puncture and Allen test not recorded.
E. Patient hyperventilation during procedure can cause false changes in
metabolic status.

33. REFFERENCE
• Basic clinical laboratory techniques- Barbara h. Estridge
• Concise book of medical laboratory technology methods and
interpretation –Ramanik Sood
• Text book of medical laboratory technology – Godkar
• Haematology - Ramanik Sood