Collection and handling of blood
VENOUS BLOOD
Specimen collection should be undertaken by specially trained phlebotomists.
Phlebotomy Tray: It is convenient to have a tray which contains all the
requirements for blood collection.
Disposable Plastic Syringes and Disposable Needles
► The needles should not be too fine, too large, or too long.
► Those of 19 or 21G are suitable for most adults.
► 23 G are suitable for children and ideally should have a short shaft (about 15
mm). It may be helpful to collect the blood by means of a winged (‘butterfly’)
needle connected to a length of plastic tubing which can be attached to the
nozzle of the syringe.
Butterfly needle
Specimen Containers :
Types of containers for hematology tests that are available commercially include:
A-Containers with anticoagulants:
▪ Containers with dipotassium, tripotassium, or disodium ethylenediaminetetra-
acetic acid (EDTA).
▪ Containers with trisodium citrate.
▪ Containers with heparin
▪ Containers with fluoride
All these are marked at a level to indicate the correct amount of blood to be
added.
B- Containers without anticoagulants (plain tubes):
which are used when serum is required.
■ Evacuated tube systems which are now in common use consist of a glass or
plastic tube/container (with or without anticoagulant) under defined vacuum,
a needle, and a needle holder which secures the needle to the tube.
■ The main advantage is that the cap can be pierced, so that it is not necessary to
remove it either to fill the tube, or subsequently to withdraw samples for analysis,
thus minimizing the risk of aerosol discharge of the contents.
■ An evacuated system is useful when multiple samples in different
anticoagulants are required.
■ The vacuum controls the amount of blood which enters the tube, ensuring an
adequate specimen for the subsequent tests and the correct proportion of
anticoagulant, when this is present.
Phlebotomy Procedure
■ The phlebotomist should first check the patient’s identity, making sure that it
corresponds to the details on the request form, and also ensure that the
phlebotomy tray contains all the required specimen containers.
■ Blood is best withdrawn from an antecubital vein or other visible veins in the
forearm by means of either an evacuated tube or a syringe.
■ It is usually recommended that the skin should be cleaned with 70% alcohol
(e.g. isopropanol) or 0.5% chlorhexidine, and allowed to dry spontaneously before
being punctured.
■ When using a tourniquet , it should be applied just above the venepuncture
site and released as soon as the blood begins to flow into the syringe or
evacuated tube – delay in releasing it leads to fluid shift and hemoconcentration
as a result of venous blood stagnation.
■ Successful venepuncture may be facilitated by :
1-Keeping the subject’s arm warm.
2-Applying a sphygmomanometer cuff to the upper arm and kept at
approximately diastolic pressure.
3-Tapping the skin over the site of the vein a few times.
■ After cleaning and drying the site and applying a tourniquet, ask the patient to
make a fist a few times.
■ If the veins are very small, a butterfly needle or 23G needle should enable at
least 2 ml of blood to be obtained satisfactorily.
■ In obese patients, it may be easier to use a vein on the dorsum of the hand,
after warming it by immersion in warm water; however, this site is not generally
recommended as vein punctures tend to bleed into surrounding tissues more
readily than at other sites.
■ Venipuncture should not be attempted over a site of scarring or hematoma.
■ If a syringe is used for blood collection, the piston of the syringe should be
withdrawn slowly and no attempt made to withdraw blood faster than the vein is
filling.
■ Anticoagulated specimens must be mixed by inverting the containers several
times.
►Haemolysis can be avoided or minimized by:
1- Withdrawing the blood slowly
2- Not using too fine needle
3- Delivering the blood gently into the collection tube.
4- Avoiding frothing during the withdrawal of the blood.
■ If the blood is drawn too slowly or inadequately mixed with the anticoagulant
some clotting may occur.
■ After collection, the containers must be firmly capped to minimize the risk of
leakage.
■ After obtaining the blood and releasing the tourniquet, remove the needle and
then press a sterile swab over the puncture site.
■ The arm should be elevated after withdrawal of the needle and pressure should
continue to be applied to the swab with the arm elevated for a minute or two
before checking that bleeding has completely ceased.
■Then cover the puncture site with a small adhesive dressing.
■ If intravenous fluids are being transfused into an arm, the blood sample should
not be collected from that arm as this is a potential source of error.
Selection of veins
Post-phlebotomy Procedure
Specimens should be sent in individual plastic bags separated from the request
forms to prevent contamination of the forms in the event of leakage.
Alternatively, the specimen tubes must be set upright in a holder or rack and
placed in a carrier together with the request forms for transport to the
laboratory.
Waste Disposal
■ Without separating the needle from the syringe, place both, together with the
used swab and any other dressings, in a puncture-resistant container, for disposal.
■ If it is essential to dispose of the needle separately it should be detached from
the syringe only with forceps or a similar tool. Alternatively, the needle can be
destroyed in situ with a special device, e.g. Sharp-X
(Biomedical Disposal Inc: www.biodisposal.com)
CAPILLARY BLOOD
Skin puncture can be used for obtaining a small amount of blood into a special
anticoagulated microcollection device. These methods are mostly used when
1- It is not possible to obtain venous blood (e.g. in infants under 1 year and in
gross obesity)
2- For point-of-care blood tests (Beside the patient).
Collection of Capillary Blood
■ Skin puncture is carried out with a needle or lancet.
■ In adults and older children, blood can be obtained from a finger; the
recommended site is the distal digit of the third or fourth finger on its palmar
surface.
■ In infants , samples can be obtained by a deep puncture of the plantar surface
of the heel .
■ The central plantar area and the posterior curvature should not be punctured in
small infants to avoid the risk of injury and possible infection to the underlying
tarsal bones, especially in newborns.
■ Clean the area with 70% alcohol (e.g. isopropanol) and allow to dry.
■ Puncture the skin to a depth of 2–3 mm with a sterile disposable lancet.
■ Wipe away the first drop of blood with dry sterile gauze.
Collect the second and following drops directly onto a reagent strip (point-of-care
blood tests) or by a 10 ml or 20 ml micropipette for immediate dispensing into
the microcollection tube.
BLOOD FILM PREPARATION
-Ideally, blood films should be made immediately the blood has been collected.
As blood samples are usually sent to the laboratory after a variable delay there
are advantages in preparing blood films when the phlebotomy is carried out.
-The phlebotomy tray might include some clean glass slides and spreaders, and
phlebotomists should be given appropriate training for film preparation.
-An automated device for making smears is also available.
-When films are not made on site they should be made in the laboratory
withoutdelay as soon as the specimens have been received.
SAMPLE HOMOGENEITY
In order to ensure even dispersal of the blood cells it is essential that specimens
are mixed effectively immediately prior to taking a sample for testing. Place the
specimen tube on a mechanical rotating mixer for at least 2 min or invert the tube
8–10 times by hand. If the specimen has been stored at 4C, it will be viscid and
the blood should be allowed to warm up to room temperature beforemixing.
SERUM
■ The difference between plasma and serum is that the latter lacks fibrinogen and
some of the coagulation factors.
■ Blood collected in order to obtain serum should be delivered into sterile tubes
with caps or commercially available plain (non-anticoagulant) evacuated
collection tubes and allowed to clot undisturbed for about 1 h at room
temperature.
■ Then loosen the clot gently from the container wall by means of a wooden stick,
or a thin plastic or glass rod. Rough handling will cause lysis. Close the tube with a
cap/stopper.
■ Some tubes contain a clot activator combined with a gel for accelerated
separation of serum, e.g. Serum separator tubes (BD Ltd).
■ The tubes, whether with or without a serum separator ,are centrifuged for 10
min at about 1200 rpm.
■ Pipette the supernatant serum into another tube and centrifugeagain for 10
min at about 1200 rpm.
■ Transfer the supernatant serum to tubes for tests or for storage.
■ For most tests, serum should be kept at 4C until used, but if testing is delayed,
serum can be stored at – 20 C for up to 3 months and at – 40 C or lower for long-
term storage.
-Frozen specimens should be thawed in a water-bath or in a 37 C incubator, then
inverted several times to ensure homogeneity before use for a test.
-Do not refreeze thawed specimens.
