UNIVERSAL PRECAUTIONS

Universal precautions may be defined as a method for controlling infection in which all blood and
certain body fluids are treated as if infected with hepatitis B, human immunodeficiency virus (HIV) or
other disease-producing blood-borne pathogens. The reason for universal precautions is that all
patients infected with blood-borne pathogens cannot be readily identified. Therefore, certain
precaution techniques are used for all patients.
Under universal precautions the following policies are applicable to laboratory personnel.

1. Skin and mucous membrane exposure to blood and other body fluids must be prevented.
2. Gloves must be worn when there is any possibility of coming in contact with blood or other body
fluids. When removing gloves they must be disposed of in biohazardous waste. Gloves must never be
reused.
3. Masks and protective eyewear (goggles) or face shields are to be worn if there is a possibility of
droplets or spattering of the blood or body fluid. Use of plexiglas shields in the work area is an
alternative.
4. Gowns or laboratory coats should be worn when working with blood or body fluids. These
coverings must be removed before leaving the laboratory and may not be taken home for washing.
5. Any skin surfaces that become contaminated with blood or body fluids should be washed
immediately. Hands must be washed upon removal of gloves.
6. It is utmost importance that any cuts or scratches be well protected from contamination with blood
or body fluids.
7. During phlebotomy, used needles should not be recapped, bent, or broken by the technologist. The
unsheathed needle should be placed directly into an appropriately labelled puncture-resistant bio-
hazard container for disposal.
8. All specimens for centrifugation must be centrifuged in a closed tube (a top must be on every tube).
9. Special care must be taken to avoid leaking of specimen tubes or containers.
10. All pipetting must be carried out using mechanical pipet devices.
11. All laboratory work benches must be decontaminated with the appropriate germicide (10% Clorox,
for example) when work has been completed (at least once per shift).
12. Instrumentation must be decontaminated prior to servicing. If this is not possible, a warning
should be placed on the equipment.
13. All materials coming in contact with blood or body fluids must be must be placed in biohazardous
waste containers when finished with and disposed of according to institution’s infective waste
disposal policy.
14. No precaution labels are used on any patient specimens because of the fact that all specimens
are considered infectious and handled accordingly.

COLLECTION OF BLOOD

The Medical Technologist most often comes in contact with a patient during the process of blood
collection. The patient in a hospital is anxious, fearful, and ill health. He is anxious about his physical
condition; he fears because he does not know what will happen next; his disease may or may not be
life threatening; and he is physically uncomfortable as a result of his illness or injury. He is also
separated from his known surroundings and family. For these reasons, a person’s mental attitude as
a often at its worst when he is in the hospital as a patient. It is important, therefore, for the medical
technologist to show the patient, at all times, the kindness and understanding that can mean so much.
When the technologist is dealing with a child, his approach is doubly important. This may be the
first time the child has had blood test. If it turns out to be a horrendous experience, it will be
remembered and feared by the child for many years. Therefore, it is important to gain the child’s
confidence before proceeding with blood collection. The child should be informed of what is going to
happen. If the child is told that the puncture will not hurt, the child’s confidence will be lost because
this statement is generally not true. A routine venipuncture may be compared to a bee sting, while the
fingerstick may be described as a mother pricking her finger with a needle or pin while sewing.
The techniques used in obtaining blood are not learned overnight. They are an art that must be
developed by study, observation, and practice, until the technologist has the necessary skill and self
confidence. Skill, patience, understanding- these are the qualities of a good phlebotomist.
Blood specimens are commonly obtained from a patient’s vein. Under some circumstances, as
outlined below under Microsample Technique, a skin puncture will be used for this purpose.
Irrespective of the method used certain techniques are common to all phlebotomy procedures:

1. The correct patient identification is critical. For hospital patients this is accomplished by checking
the identification wrist band for them correct name and hospital identification number. The procedure
for out-patients is not as easy. The phlebotomist should ask the patient for his or her full name and
any other information specific for that patient that can be verified y the requisition slip.
(Misidentification of a patient is a serious error and can have disastrous implications for the patient).
2. The correct specimen identification is equally as important as patient identification. Each blood
specimen obtained should be labelled with the patient’s first and last name, the hospital identification
number, patient location, time, date, and the phlebotomist’s initials.
3. To be consistent with laboratory safety guidelines and universal precautions, gloves must be worn
at all times will performing phlebotomy techniques to protect the technologist from acquiring blood-
borne infections such as hepatitis B, or HIV. In addition, the phlebotomist’s hands should be washed
between each patient when removing gloves.
4. The puncture site should be cleaned by rubbing vigorously with a pad thoroughly moistened with
70% ispropanol (v/v). The area is then dried using sterile gauze. Once the phlebotomy site has been
cleaned the decontaminated area should not be touched. Betadine should not be routinely used to
clean phlebotomy area because contamination with this substance will cause some erroneous test
results (falsely elevates potassium, uric acid, and phosphorus results).
5. All sharp objects such as lancets and needles must be disposed of in special puncture-resistant
disposable needle containers labelled as biohazardous. Needles should not be bent, broken, or
resheathed before disposal. All other objects such as gauze and alcohol prep swabs should be
placed in biohazardous wasted containers.

COAGULATION TESTING REQUIREMENTS

Coagulation specimens require special attention. Poor techniques used in collection,

processing, or storage will cause misleading and inaccurate results

Specimen Collection

1. A clean venipuncture is absolutely necessary. Any contamination of the blood with with tissue
fluid leads to incorrect results. (a) for routine coagulation procedures, the vacutainer blood
collection system is adequate. When this system is used, the tube for coagulation studies should
be the second or tube filled. If only coagulation tests are ordered, a plain tube should be filled with
2 to 3mL of blood first and then the coagulation tube filled. The first tube may then be discarded
upon returning to the laboratory. (b) When obtaining a blood specimen for special coagulation
procedures, a two-syringe technique may be applied using plastic syringes. Withdraw
approximately 2mL of blood into the first syringe. Quickly and carefully disconnect this syringe
from the needle (leaving the needle in the patient’s vein) and connect the second syringe to the
needle. Proceed with the venipuncture, discarding the blood in the first syringe. (Prior to
performing the two-syringe venipuncture, make certain the first syringe is easily removed from the
needle but fits snugly enough so that blood will not leak out at the connection.) (c) Capillary
specimens are generally not used for coagulation testing because of the difficulty in obtaining a
blood specimen without trauma and free of tissue juice.
2. When phlebotomizing the patient it is important to release the tourniquet as soon as the blood
enters the first tube (or the syringe, if used). When using vacutainer tubes care must be taken to
allow the tube to fill until there is no vacuum remaining. The anticoagulant-to-blood ratio is
important in coagulation studies. The tube must be filled to within ±10% of its expected volume.
Greater of lesser amounts of blood will yield incorrect coagulation test results.

3. All test tubes used for coagulation studies should have a “noncontact” surface. That is, the inside
surface of the tubes should not be of a material (such as glass or soda lime) that will react or
activate the coagulation factors. Most vacutainer tubes for collecting coagulation studies have a
siliconized surface.

4. The anticoagulation of choice for coagulation studies is 0.109 M sodium citrate (buffered or
nonbuffered). Alternatively, 0.129 M sodium citrate may be used, but is not the recommended
concentration. The ratio of blood to anticoagulant is 9 parts blood to 1 part sodium citrate. It is
critical that the proper amount of blood (±10%) be added to the anticoagulant. The presence of a
buffer in the anticoagulant maintains a more stable pH.

5. Blood specimens with a high hematocrit will contain less plasma in relation to the volume of blood
in the tube. As a result, when the blood has been centrifuged, the plasma fraction will contain an
increased concentration of anticoagulant (sodium citrate). (The 0.5mL of anticoagulant will be
diluted with plasma.) during testing, as calcium is added to the test mixture, it may combine with
the excess anticoagulant present instead of being available for the coagulation reaction.
Therefore, whenever a patient’s hematocrit is 55% or higher the amount of sodium citrate in the
collection tube should be decreased (or the amount of anticoagulant may be held constant, and
the volume of blood collected increased). The following formula may be used to determine the
proper amount of sodium citrate.

Amount of sodium citrate

100 - Hematocrit
= _______________ x mL of whole blood used

595 - Hematocrit
For example, when a patient has a hematocrit of 60%. 0.34mL of 0.109 M sodium citrate should be
placed in the tube and 4.5mL of whole blood added. When altering the amount of anticoagulant (or
specimen) the sample must be obtained using a syringe.

Specimen Processing

1. Each blood specimen should be checked for clots prior to testing. Because the tubes must be
centrifuged with their tops on, the red cell cell layer may be checked for clots after the plasma has
been removed. The presence of a clot irrespective of how small it is renders the specimen
unacceptable for coagulation testing.
2. Unless otherwise noted, the coagulation specimen should be centrifuged with 1 hour of obtaining
the sample. It is desirable to complete testing within 2 to 4 hours, depending on the specific
procedure.
3. All pipets, test tubes, and reaction cups which come in contact with the test plasma should have a
“noncontact” surface (e.g., plastic, siliconized).
4. The plasma specimen should be in a stoppered container at all times, except when testing. This
prevents loss of CO2, and a resultant pH change of the plasma.
5. For most coagulation procedures, unless noted under specimen requirements, if the specimen
will be tested within 2 hours of collection it may be kept at room temperature. If there is to be a
delay beyond 2 hours it is advisable to keep the specimen on ice. Exceptions to this rule are
specimens for prothrombin time, factor VII assay, and platelet function studies. These specimens
should be maintained at room temperature. Factor VII may become activated by the cold. Also,
plasma for the euglobin clot lysis procedure must be tested or frozen within 30 minutes of
collection.

6. With the exception of platelet function studies, plasma for coagulation testing should be platelet
poor (plasma platelet count less than 15,000/µL). Centrifugation for 15 minutes at 2500 x g will
routinely produce acceptable platelet-poor plasma. Use of commercially available serum-plasma
separators (Sure-Sep, Organon Teknika, Durhma, NC) produces excellent quality platelet-poor
plasma acceptable for coagulation testing. In addition, several small table top high speed
centrifuges are available for more rapid centrifugation of plasma samples. Platelet-poor plasma
may be obtained using speeds of 1300 x g for 2 minutes. These centrifuges cost less than $2000
and are excellent for rapid turnaround of test results when necessary
7. An alternative method to crushed ice for keeping specimens cold is the commercially available
Kryorack (Streck Laboratories, Inc., Omaha, Nebraska). This test tube holder contains an
aqueous solution within a sealed unit and is available in various sizes. When placed at freezer
temperatures (-18°C) for 8 hours, it maintains a temperature for the test tubes of less than 8°C for
8 hours at room temperature. The tubes do not come in contact with water or ice, affording a
more convenient method of refrigerating plasma.

8. Hemolyzed plasma is indicative of a traumatic venipuncture and should not be used for
coagulation studies. Results will not be dependable. Also, automated instruments may not be
able to detect the end point on some lipemic or icteric plasma samples.

Plasma Storage

1. Platelet-poor plasma may be stored at -40°C or lower for at least several weeks without loss of
most factors. The plasma should initially be frozen (in covered plastic containers) very quickly
using liquid nitrogen or temperatures at -80°C to -40°C.
2. When thawing frozen plasma it should be done rapidly in a 37°C incubator or water bath. Remove
the plasma from the incubator as soon as it is thawed. Plasma cannot be refrozen.

Coagulation Testing

1. Most coagulation studies are carried out at 37°C. It should be noted that specimens incubated in
dry heat take slightly longer to reach 37°C than those incubated in a water bath. It is essential,
when required, that the specimens and reagents reach the proper temperature of 37°C before
proceeding with the test. Overheating or prolonged heating at 37°C, however, may lead to
destruction of some of the coagulation factors and, therefore, a prolonged clotting time. The
temperature of the incubator should not fluctuate more than ±0.5°C..
2. Accurate timing of clotting times is extremely important. Many of the procedures are timed within
one-tenth of a second, so that the initial starting and stopping of the stopwatch must be done
precisely.
3. There are four general techniques in widespread use for reading the end point of clotting
procedures. The tilt tube method requires gentle tilting of the tube back and forth at the rate of
about once per second until a fibrin web is formed. The nichrome wire loop technique employs
the use of a wire loop that is passed through the mixture at the rate of two sweeps per second
 until a formed clot adheres to the loop. (In these first two techniques, a light source without glare
is important. A black background also facilitates the end point readings.) The third and fourth
procedures employ the use of automation: a clot is detected either by use of a moving probe
immersed in the mixture that is triggered by clot formation or by the change in optical density of
the mixture when a clot forms.
4. The appearance of the clot formed frequently depends on its rate of formation. Normal, or only
slightly prolonged clotting times show a pronounced clouding of the mixture that is easy to see
with the human eye. Prolonged clotting times, however, form more slowly and may first appear as
very fine fibrin threads. The mixture becomes cloudy (opaque) very slowly; it is often difficult to
detect when final end point occurs. Automated coagulation analyzers based on optical density
should be able to read this latter type of clot formation.
5. Historically, almost all coagulation procedures have been performed in duplicate and the two
numbers averaged for a final result. This is time consuming and expensive. Because of more
accurate and precise pipets and instrumentation now in use, this double testing is becoming
unnecessary. Each laboratory should check the accuracy and precision of their methods in an
effort to change over to single sample testing. Alternatively, sample and reagent volumes may be
reduced on some instrumentation so that duplicates may be performed using the same
plasma/reagent volumes as for a single sample.