VACUUM BLOOD
COLLECTION
Terry Kotrla, MS, MT(ASCP)BB
Revised with permission January 2016 by Lois Wagoner, MT(ASCP)
Introduction
• The vacuum blood collection system consists of
a double-pointed needle, a plastic holder or
adapter, and a series of vacuum tubes with
rubber stoppers of various colors.
• The evacuated tube collection system will
produce the best blood samples for analysis.
• The blood goes directly from the patient vein into
the appropriate test tube.
Multi-Sample Needle
• The bevel is the slanted opening at the end of the needle.
• Needle length (shaft) ranges from 1 to 1 ½ inches.
• Threaded hub screws into needle holder
• The rubber sheath makes it possible to draw several tubes of
blood by preventing leakage of blood as tubes are changed.
Bevel
• Bevel is slanted opening at
end of needle.
• Needle must be oriented so
that bevel faces up prior to
insertion.
Needle Gauge
• The gauge of a needle is a number
that indicates the diameter of its
lumen.
• The lumen, also called the bore, is the
circular hollow space inside the
needle.
• The higher the gauge, the smaller the
lumen.
• The most frequently used gauges for
phlebotomy are 20, 21 and 22
Holder
• The holder for vacuum blood collection
is a plastic sleeve into which the
phlebotomist screws the double pointed
needle.
• The most current guidelines require
that all holders are for single use only.
Vacuum Collection Tubes
• Vacuum collection tubes are sealed with a partial vacuum inside by
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rubber stoppers.
The air pressure inside the tube is negative, less than the normal
environment, which creates the vacuum in the tube.
After inserting the longer needle into the vein, the phlebotomist
pushes the tube into the holder so that the shorter needle pierces the
stopper.
The difference in pressure between the inside of the tube and the vein
pulls the blood into the tube, which allows the tube to fill with blood.
The tubes are available in various sizes for adult and pediatric
phlebotomies
Plastic tubes have replaced most glass tubes for safety reasons
Additives
• Found in almost all blood collection tubes used today
• Most common additive is an anticoagulant which
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prevents clotting of the blood
Some tubes have a clot activator to help the clotting
process start and finish completely
A preservative is present in some tubes to help maintain
sample stability
The polymer gel found in some tubes will move between
the cells and liquid portion of the sample during
centrifugation to form a physical barrier
The color of the tube stopper indicates what additive, if
any, the tube contains.
Anticoagulants
• Tubes containing anticoagulants have the precise
amount needed to mix with the amount of blood
that will fill the tube.
• It is important to completely fill each tube to its
intended volume so that the proportion of blood to
chemical additive is correct; otherwise, the test
results may not be accurate or the specimen will
be rejected and will need to be recollected.
• It is critical to thoroughly mix the blood with the
anticoagulant by gentle inversion so that it can
work properly and produce the intended
specimen.
Blood Cultures
• Is a special collection to detect bacteria growing
in the blood.
• Blood may be collected into a Yellow stopper tube
containing Sodium Polyanethol Sulfate (SPS) or
may be collected into Blood Culture Bottles.
• Site preparation VERY important.
• Will be covered later in a separate lab activity.
(Light) Blue Stopper
• Anticoagulant: Sodium Citrate
• Ratio of blood to anticoagulant is 9:1
• Tests include Coagulation studies such as PT,
PTT and fibrinogen
• MUST BE FILLED COMPLETELY!!! NO
EXCEPTIONS
• http://www.austincc.edu/kotrla/phb_ltblue
Red Stopper
• No additive in glass tube
• Clot activator in plastic tube
• No anticoagulant present
• Tests using serum which include: most blood
chemistries, AIDS antibody, viral studies, serology
tests, Blood Bank testing using serum.
• http://www.austincc.edu/kotrla/phb_red
Serum Separator Tubes (SST)
• SST = Serum Separator Tube
• Can be red/black mottled, gold or red with black stopper.
• Gel forms physical barrier during centrifugation that
separates serum from cells permanently
• All tests using serum except Blood Bank
• http://tinyurl.com/8jznm
Green Stopper
• One of the following formulations:
• sodium heparin
• lithium heparin
• ammonium heparin
• Used for STAT blood chemistries utilizing plasma.
• http://www.austincc.edu/kotrla/phb_green
Green Plasma Separator Tube
• Plasma Separator Tube=PST
• Anticoagulant is heparin, so it can be centrifuged
immediately.
• Gel forms physical barrier during centrifugation,
permanently separates plasma from red blood cells
Lavender Stopper
EDTA (ethylenediaminetetraacetic)
• Used for Hematology studies: CBC, WBC count,
Hemoglobin, Hematocrit, Platelet count,
Reticulocyte count, differential.
• http://www.austincc.edu/kotrla/phb_purple
• Anticoagulant =
Pink Stopper
• Primary use is for blood bank testing using the gel
system, which requires plasma.
• Anticoagulant = EDTA (ethylenediaminetetraacetic)
• May also be used for hematology if it has not been
centrifuged.
Gray Stopper
• Additive (read label):
• Potassium oxalate and sodium fluoride (plasma)
• Sodium EDTA and sodium fluoride (plasma)
• Sodium fluoride (serum)
• Test include: Glucose, Blood Alcohol (ethanol) levels,
lactic acid
• Sodium Fluoride is a preservative that prevents glycolysis
• http://www.austincc.edu/kotrla/phb_gray
Order of the Draw
• It is critical that the order of the draw is followed to
minimize the effect of carryover
• Carryover of additive from one tube to the next can
adversely affect test results
• Each lab should document it’s order of the draw
• The Clinical and Laboratory Standard’s Institute (CLSI)
has developed an order of the draw that all phlebotomist
should know
CLSI recommended Order of the Draw
1. Sterile/Blood cultures or Yellow SPS tube
2. Coagulation tube (Sodium Citrate) Light Blue
3. Serum tube (with or without gel) Red, Gold, Mottled
4. Other additives
a. Heparin (with or without gel) Green
b. EDTA - Lavender/Pink
c.
Glycolytic Inhibition tube - Gray
Sodium fluoride & potassium oxalate or
Sodium fluoride & sodium EDTA or
Sodium fluoride only
Specialty Tubes
• The following tubes are used less frequently.
• Your clinical site may use these and you need to be aware
of the additive and uses.
• The order in which these tubes are drawn is usually
based on the additive in the tube, but may vary from lab to
lab
• It is the phlebotomist responsibility to learn about any
tubes that may be used by their lab and the correct
placement of those tubes in the order of the draw
Black Stopper
• Buffered Sodium Citrate
• Only used for Westergren sedimentation rate
determination
• MUST BE FILLED COMPLETELY!!!
NO EXCEPTIONS!!!
Royal Blue Stopper
• Color of tube label indicates additive, if any:
• purple - EDTA
• green - heparin
• red – none
• Order of the draw will be determined by additive present.
• Used for trace metal analysis, nutrients and toxicology studies such
as: Antimony Arsenic, Cadmium, Calcium, Chromium, Copper, Iron,
Lead, Magnesium, Manganese, Zinc
Tan Stopper
• Anticoagulant: K2EDTA
• Specifically for lead analysis although royal blue
can be used.
Yellow Stopper
• Sodium polyanethol sulfonate (SPS)
• SPS for blood culture specimen collections in
microbiology.
• Tube inversions prevent clotting.
• Acid citrate dextrose additives (ACD)
• ACD for use in blood bank studies, HLA phenotyping,
DNA and paternity testing.
Blood Composition
• As blood flows through the body it is made up of plasma
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and cellular elements.
The cellular elements include red blood cells, white blood
cells and platelets.
Plasma in the body is composed of water and dissolved
substances, such as proteins, nutrients, carbohydrates,
lipids, minerals, gases, vitamins, hormones, antibodies,
fibrinogen and waste products.
The lab test for these different substances and cellular
elements require different types of blood specimens.
The 3 types of blood specimens collected by venipuncture
are serum, plasma, and whole blood
Types of Blood Specimens: Serum
• Serum is the liquid portion of a blood specimen that has been allowed
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to clot
Serum samples must be allowed to sit upright for at least 30 minutes
to allow the clotting process to occur and reach completion
The sample must then be spun down, which allows the serum to rise
to the top of the sample as the clot moves to the bottom
Serum does not contain any coagulation factors
Many chemistry and serology test are preformed on serum
The most common tubes that produce serum include red, gold,
red/black mottled and tiger-top tubes
Other tubes that produce serum but are not used as often include
royal blue with a red label and grey with only sodium fluoride
Types of Blood Specimens: Plasma
• Plasma is the clear liquid portion of an anticoagulated blood specimen
• Anticoagulated specimens must be mixed well immediately after
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drawing so the anticoagulant is evenly dispersed in the sample.
Anticoagulated samples may be spun down as soon as they arrive in
the lab. This makes them an excellent choice for STAT lab test.
Plasma contains all of the coagulation factors except one. Different
anticoagulants inhibit or interfere with different coagulation factors.
Plasma is used for a variety of test, including coagulation studies, stat
chemistries, and blood banking.
The most common tubes that yield plasma are light blue, green, pink
and grey stopped tubes which include an anticoagulant.
Other tubes that produce plasma include royal blue with a green or
lavender label.
Types of Blood Specimens: Whole Blood
• Whole Blood is an anticoagulated specimen in which the cellular
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elements remain suspended in the plasma for testing purposes.
Whole Blood samples are never centrifuged; they are often placed on
a mechanical rocker to keep the cellular elements suspended until
testing can be preformed.
Whole blood samples are used for Hematology test, Blood cultures,
Genetic and Tissue testing.
The most commonly drawn tube for whole blood testing is the
lavender topped tube used for Hematology test.
Other whole blood samples are collected in yellow (both SPS and
ADC), black, tan, and royal blue with a purple label.
Patient Identification
• It is vitally important that the phlebotomist correctly identifies the
patient.
• Do not offer the patient a name to respond to. Always ask the
patient to state their name.
• All hospitalized patients have an identification arm band with their
name, hospital identification number and other pertinent information.
• Always compare the laboratory test request slip name and ID number with the
name and ID number on the patient's hospital arm band AND with the name the
patient states for you.
• If there is any discrepancy, do not draw the patient's blood.
• For an out-patient – site specific protocols must be followed which
may include:
• Verify the patient's identity by having the patient give you additional identifying
information such as a unique ID number, date of birth or address.
• Patient may be asked to review and initial label.
Preparation
• Wash or disinfect your hands and put on gloves
• Introduce yourself, state your mission
• Properly identify the patient
• Ask the patient if they have had any problems during
previous blood draws
• If they have had previous problems, discuss these further
to help you prepare an appropriate plan for the patient
• If the patient has not had a previous blood draw, explain
the procedure to them
• Choose the appropriate tubes for the tests requested
Tourniquet Application
• Apply approximately 3-5 inches above antecubital fossa.
• If the skin appears blanched above and below the
tourniquet it is too tight.
• If your finger can be inserted between the tourniquet and
the patient's skin it is too loose.
Palpate
• After tourniquet application have patient clench fist.
• Feel for a vein that rebounds (bounces) when pushed or tapped on.
• PALPATE potential veins to help determine the following attributes:
size
• direction
• depth
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• A slight rotation of the arm or wrist may help to better expose a vein
that may otherwise be hidden.
• Use your index finger and middle finger to palpate.
• Never use your thumb to palpate a vein.
Vein Selection
• Choose the vein that is large and accessible.
• Choose the vein that feels the best, which may or may not
be the most visible.
• Avoid bruised and scarred areas.
• Avoid veins which are sclerosed or occluded.
• Palpate the vein above and the below the chosen
venipuncture site to note the depth, diameter and
direction of the vein
Can’t Feel the Vein?
• Tricks to Help Distend Veins:
• Make sure the tourniquet is not loose
• Rotate the patient’s wrist
• Ask the patient to make a fist.
• Do not allow the patient to pump their fist, this can lead to erroneous
test results.
• Hand the patient a couple of plastic tubes to grasp, as this is sometimes
easier for the patient.
• Have the patient dangle arm below the heart level for 1-3 minutes.
• Warm the area with a hot pack or warm, moist cloth heated to
approximately 42°C.
• Remember to not leave the tourniquet on for more than 1 minute.
• If you are unable to locate a usable vein consult an experienced
phlebotomist for assistance and guidance.
Veins used for drawing blood
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Median cubital vein - first choice, well supported,
least apt to roll
Cephalic vein - second choice
Basilic vein - third choice, often the most prominent
vein, but it tends to roll easily and makes venipuncture
difficult
Median Cubital – First Choice
• This vein is located in the antecubital fossa (the area of
the arm in front of the elbow)
• Well anchored vein, usually large and prominent.
• Very few problems. Offering the best chance for a close to
painless puncture, as there are few nerve endings close
to this vein.
Vein Selection
Cephalic Vein-Second Choice
• Cephalic vein which is located on the upper or shoulder
side of the arm.
• This vein is usually well anchored.
• The cephalic vein may lie close to the surface. A low
angle of needle insertion must be used to avoid possible
spurting or blood forming a drop at the puncture site. (15°)
Veins for Venipuncture
• These are NOT listed in the order of preference but
illustrates the usual position of the veins.
Basilic Vein-Third Choice
• Located on the under side of the arm (pinky side).
• In many patients this vein may not be well anchored and
will roll, making it difficult to access with the needle.
• Syringe draw should be considered as it gives the
phlebotomist more control over a rolling vein.
• Pooling of blood and hematoma formation possible.
• Exercise caution when drawing from this area.
• The basilic vein is close to the brachial artery so there is more risk
of hitting an artery.
• The basilic vein lies close to the brachial nerve which may result in
injury to the nerve.
• This area is often more sensitive, thus may be slightly more painful
for the patient
Cleansing the Site
• After selecting a vein, remove the tourniquet.
• Clean the selected site with a prepackage 70% isopropyl alcohol swab.
• Start at the center of the selected site and rub the alcohol swab in a circular
motion moving outward from the site. Use enough pressure to remove all
perspiration and dirt from the puncture site.
• Discreetly look at the swab when finished; if it appears excessively dirty
repeat the cleansing process with a fresh alcohol swab.
• After cleansing do not touch the site, if the vein must be repalpated the area
must be cleansed again.
Assemble Equipment
• After cleaning the site, assemble the equipment. This will
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allow the site time to dry.
Twist needle into holder.
Select appropriate tubes and insert first tube into holder.
Reapply the tourniquet
DO NOT remove cap until right before you are ready to
draw.
Re-Apply Tourniquet and Prepare to Draw
Anchoring the Vein
• Using your non-dominant hand, position your thumb
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below the intended puncture site and pull the skin down
and away from the intended site until the skin is taut
You may have to pull slightly off-center so that your thumb
is not in the way of the holder
Use the rest of your fingers of your non-dominant hand to
hold the patient’s arm steady
By pulling the skin taut, the puncture is more likely to be
less painful to the patient
Large veins that are not well anchored in tissue frequently
roll. Use extra care to anchor these veins for a successful
venipuncture.
Performing the Draw
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Make sure the tourniquet has been properly reapplied.
Uncap the needle using your non-dominant hand
Briefly inspect the needle for defects
Hold the prepared holder with the bevel of the needle facing up.
Use the thumb of the non-dominant hand below the puncture site to
anchor the vein and pull the skin taut.
• The needle entering the site should not touch the thumb of the
phlebotomist.
• Position the needle in the same direction as the vein, enter the skin
and penetrate the vein at a 15-30 degree angle in one swift, smooth
motion to decrease the patient's discomfort.
• If you enter too slowly
• Blood may leak out at the puncture site creating a biological hazard
• Your view of the puncture site may be obstructed by the pool of blood
• The patient usually feels more pain and discomfort from the needle entry
Performing the Draw (continued)
• The bevel of the needle should enter and remain in the
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center of the vein. You may feel a slight “give” or decrease
of resistance when you are in the vein.
Using your non-dominant hand, carefully push the first
tube onto the back of the needle in the holder and allow
the first tube to fill.
Stabilize the needle and use counter pressure when
putting tubes on or off the needle to insure that the needle
does not move.
Change tubes as needed until all tubes have been filled.
Remove the tourniquet when your last tube is filling or
within one minute of application, which ever comes first
Performing the Draw
Ending Draw - Release Tourniquet
• Tourniquet cannot be in place more than 1 minute.
• Release the tourniquet as the last tube is filling.
• Use one handed method of release.
Ending Draw -TTN
• Release Tourniquet
• Release last Tube from needle.
• Hold gauze sponge or biowipe above needle without
applying pressure to the puncture site.
• Swiftly withdraw Needle.
• As soon as needle is withdrawn
• Apply pressure to puncture site with your non-dominant hand.
• Immediately activate the needle safety device according to the
manufacture’s directions and discard into sharps container
• If possible, have patient continue to apply pressure.
Ending the Draw TTN
Activating Safety Device
• NEVER take your eyes off the needle until the
safety device is activated.
• Two hands – one applies pressure to site after
needle is removed, the other is used to activate
the device. DO NOT remove hand holding
pressure on site until safety device is activated.
• DO NOT USE YOUR OTHER HAND TO SNAP
DEVICE INTO PLACE…EVER!
BD Eclipse
• The BD Vacutainer® Eclipse™ Blood Collection Needle is a
safety-engineered multi-sample blood collection needle.
• It features a patented safety shield that allows for one-handed
activation to cover the needle immediately upon withdrawal
from the vein and confirms proper activation with an audible
click
• Look CLOSELY at the position of the thumb. DO NOT go
higher with your finger as this may lead to a needle stick injury.
Needle Disposal
• As soon as needle safety device is activated
immediately dispose of entire assembly in a
biohazard sharps container.
• CAUTION: Never attempt to shove device into a
full sharps container. This may lead to a needle
stick injury if your finger slips inside the holder
and your finger may be pierced by back end of
needle.
Mixing and Labeling Tubes
• Gently invert all tubes multiple times to insure proper
mixing of the blood and additive
• Label all tubes appropriately at the patient’s side.
• NEVER take unlabeled tubes from the patient’s presence.
• Minimum information:
• Patient’s full name, last name first
• ID number (or Date of Birth)
• Date, time and your initials
• Label the tubes from the stopper downward
Checking Site
• Gently remove gauze or biowipe.
• Inspect area for continued bleeding or swelling.
• If patient is still bleeding DO NOT leave, continue to apply pressure
• Recheck the site every minute
• Sometimes it is helpful to have patient elevate arm while applying
pressure, this slows blood flow to the area.
• Once bleeding has stopped place bandage or Coflex over
site.
• Some patients are allergic to bandages
• Coflex is wrapped over gauze and around the arm
• It sticks to itself and provides slight compression
• Tell patient to remove in 10-15 minutes.
Leaving
• Discard all used materials
• hint- place all wrappers, alcohol swab, needle cap in palm of
gloved hand, remove glove.
• Thank patient.
• Wash or sanitize hands.
• Leave
Problems with Needle Insertion
Bevel against vein wall.
Collapsed vein.
Swelling at site, hematoma,
immediately withdraw needle.
Needle not in vein, move forward.
Problems with Needle Insertion
Needle inserted too far, back up.
Needle inserted into artery,
IMMEDIATELY withdraw needle.
Safety Devices
• http://tinyurl.com/9bovf safety device animation
Sources of Error
Failure to insert the needle completely into the vein.
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Puncturing the stopper before entering the vein.
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The phlebotomist should feel resistance initially following insertion of the
needle, the resistance is almost immediately followed by a sensation of free or
easier movement as the needle enters the vein.
With experience you will feel a “pop” or “give” as the needle enters the vein.
If the phlebotomist partially pushes the evacuated tube onto the needle before
inserting the needle into the vein, there is a risk of puncturing the stopper and
releasing the vacuum.
If after pushing the tube onto the back end of the needle once the needle is in
the vein there is no blood change tubes to see if the problem is a defective
tube.
Not anchoring the vein before inserting the needle. The vein must
be held in place for successful needle penetration.
"Bouncing" the needle on the skin before guiding it into the vein.
This results in contamination of the needle and it should be
discarded.
Not keeping the holder stationary during tube change. This may
cause the needle to go through the vein when pushing the blood
collection tube onto the back end of the needle OR cause needle
to come out of vein during tube removal.
Rejection of Samples
1. Hemolysis - this is usually caused by a procedural error such as using too
small of a needle, or pulling back to hard on the plunger of a syringe used for
collecting the sample. The red cells rupture resulting in hemoglobin being
released into the serum or plasma, making the sample unsuitable for many
laboratory tests. The serum or plasma will appear red instead of straw
colored.
2. Clotted - failure to mix or inadequate mixing of samples collected into an
additive tube. The red cells clump together making the sample unsuitable for
testing.
3. Insufficient sample (QNS) - certain additive tubes must be filled completely.
Incorrect blood to additive ratio will adversely affect the laboratory test results.
When many tests are ordered on the same tube be sure to know the amount
of sample needed for each test.
4. Wrong tube collected for test ordered. Always refer to procedure manual
when uncertain as to which tube is required for the test ordered.
5. Improper storage - certain tests must be collected and placed in ice,
protected from light or be kept warm after collection.
6. Improperly labeled – There are strict guidelines for labeling. Failure to
correctly label a sample will result in the sample being rejected.
First Aid Following Needle Stick Injury
• Be careful not to stick yourself with a used
needle.
• If an accidental stick does occur immediately
 Go to the sink, turn on the water, and bleed the site
well by alternating squeezing and releasing the area
around the site.
 Do this for approximately 3 to 5 minutes.
 Afterwards scrub the site with an alcohol swab.
 Follow with a thorough hand washing.
Report it to your instructor immediately.
The End
• Revised August 29, 2013
• Revised with permission by Lois Wagoner, MT (ASCP) January 22, 2016