SOP’s of Phlebotomy

Phlebotomy, the drawing of blood, has been practiced for centuries and remains one of the Most common invasive procedures in health care. However, practice varies considerably between countries, and between institutions and individuals within the same country. These differences include variations in blood sampling technique, training (both formal and “on the job”), use of safety devices, disposal methods, device reuse, and availability of hepatitis B vaccine.

Purpose of Phlebotomy:

To obtain blood samples from patients needed for laboratory testing, with proper specimen identification and handling, while ensuring patient and staff safety

Materials and Equipments Required:

• Supportive cushion
• Specimen collection tubes
• Disposable gloves
• Ethanol/Isopropyl alcohol
• Tourniquet
• Disposable syringe
• Cotton balls
• Waste disposal bin
• Labeling sticker/ Marking pen
• Needle Destroyer

Procedure:

Do	Do Not
DO carry out hand hygiene (use soap and water or alcohol rub), and wash carefully, including wrists and spaces between the fingers for at least 30 seconds.	DO NOT forget to clean your hands
DO use one pair of non-sterile gloves per procedure or patient	DO NOT use the same pair of gloves for more than one patient DO NOT wash gloves for reuse
DO use a single-use device for blood sampling and drawing	DO NOT use a syringe, needle or lancet for more than one patient
DO disinfect the skin at the venepuncture site	DO NOT touch the puncture site after disinfecting it
DO discard the used device (a needle and syringe is a single unit) immediately into a robust sharps container	DO NOT leave an unprotected needle lying outside the sharps container
Where recapping of a needle is unavoidable, DO use the one-hand scoop technique	DO NOT recap a needle using both hands
DO seal the sharps container with a tamper-proof lid	DO NOT overfill or decant a sharps container
DO place laboratory sample tubes in a sturdy rack before injecting into the rubber stopper	DO NOT inject into a laboratory tube while holding it with the other hand
DO immediately report any incident or accident linked to a needle or sharp injury, and seek assistance; start PEP as soon as possible, following protocols (PEP: post-exposure prophylaxis)	DO NOT delay PEP after exposure to potentially contaminated material; beyond 72 hours, PEP is NOT effective

1. Venepuncture:

• Taking blood from paediatric subjects may be physically traumatic for the infant or child and emotionally traumatic for the mothers. So if sufficient blood is not obtained on the first attempt, a second attempt may be made. If the second attempt also fails, a second person may attempt to obtain the specimen. No more than three attempts should be made for any child at a time. If blood is not obtained after three attempts, the child may be scheduled to return on a later for additional attempts by the most experienced staff member.
• If three attempts must be made by fieldworker he/she should leave one venepuncture site undisturbed and informs the clinician who will attempt thrice on a peripheral vein if this is not successful, clinician proceeds to a femoral prick.
• FEMORAL PRICKS: only performed by clinicians in older children/adults, femoral area needs very thorough cleaning and sterile gloves are donned before the prick of the vein located medial to the artery.

BLOOD TAKING PROCEDURE:

1. Confirm identity of patient
2. Explain the procedure to patient/guardian. Let the patient be on the mother’s/guardian’s lap or comfortable on an examination coach/bed or seated on a chair. Ask for assistance in restraining a child if necessary BEFORE you begin.
3. Wash your hands thoroughly before the procedure; dry your hands with clean hand towels or gauze.
4. Ensure you have all the equipment that you need. Label containers with patient initials/name, screening/patient number, date, time, ward, type of sample.
5. Arrange specimen containers in the order they will be used.
6. If blood culture is required:
   • Ensure that BACTEC bottle is not expired.
   • Remove the “bottle cap”, clean the rubber top with sterile alcohol swab and allow to dry.
7. Apply the tourniquet on the mid-upper arm or just above the wrist depending on whether you intend to use veins in front of the elbow or back of the hand, respectively.
8. Wear clean gloves.
9. Identify the veins to approach.
10. Cleanse the area. Use alcohol swabs, cleansing from the venepuncture site outwards, 3 or more times. Allow the alcohol to dry. DO NOT wipe or blow on it to dry. Wash with soap and water if visibly dirty prior to swabbing with alcohol.
11. Assemble venepuncture system using aseptic technique.
12. Perform the venepuncture. Prick the vein gently, if in correct place you will notice a “flash of blood” then slowly pull the syringe plunger to collect specified amount of blood.
13. Release the tourniquet.
14. Collect appropriate volumes of samples in a sterile syringe or by connecting the blood collection tubes to the needle adaptor (volumes are specified in your study/area of work blood sample collection SOP).
15. Withdraw the needle and apply a dry swab. Ask the patient or parent/guardian to hold the swab in place for a few seconds or fold elbow to hold swab in place until bleeding stops usually in a few minutes. If bleeding continues leave swab in place and report to a clinician.
16. If a blood culture is required, place required volume in blood culture bottle first (reduce risk of contamination) before distributing in other bottles. If you are using a scalp needle (butterfly) and syringe, disconnect the butterfly tube and connect a 21-gauge needle to the syringe. Pierce through the rubber top of the culture bottle (cleaned per step 6b) making sure to control the suction effect on the syringe to place required volume.
17. Dispense appropriate volumes to respective tubes. Do not put the syringe down before all the samples have been dispensed and avoid touching the inside of the tubes with the syringe.
18. Make sure the phlebotomist’s initials, date and time of sample collection are marked clearly on the containers and laboratory request form.
19. Dispose of all clinical/laboratory waste into the appropriate containers.
20. Clean the working place, leave clean for next use. Label sample with patient’s name, IP/OPD number, date and time of collection and indicate your initials on the sample collected and the request form.
21. You may need to log in details in a specified log book.
22. Thank parent or guardian for their cooperation.
23. Hand over the samples to the laboratory(s) without delay.

2. FINGER PRICK FOR CAPILLARY SAMPLE:

Ensure that you have all the things that you will need BEFORE beginning the procedure. These are similar to those for blood sample collection but sterile lancets will also be required.
Labelling of slides: Label the slide in pencil on the frosted portion of the slide according to the study protocol involved. Make sure the slide is free from grease and if traces of oil can be observed on the slide.

Preparation of Blood Smears

1. Put on a clean pair of gloves.
2. Clean the middle or ring finger with cotton swab soaked in 70% alcohol.
3. Puncture the ball of the finger with a sterile lancet. DO NOT REUSE LANCETS!!!
4. Wipe out the first drop of blood and apply gentle pressure on the finger to release a free flowing drop of blood
5. For the thick smear, collect the drop of blood by touching the drop slightly, controlling the amount of blood to be collected (approx10-12 ul of blood)
6. Place the slide on the slide template provided and spread the drop using the edge of another clean slide avoiding formation of bubbles.
7. For the thin smear, collect a drop of blood by touching the drop slightly to acquire approx. 2-4 ul of blood.
8. Place the slide on the slide template provided
9. Using a spreader with a smooth edge, touch the drop of blood with the spreader and allow blood to spread along the spreader without touching the edges of the slide.
10. Raise the spreader to an angle of 45º and spread at moderate speed in order to create a feathered edge of the thin smear before reaching the other end of the slide.
11. Protect the smear from flies and dust as you prepare to transport it to the lab.
12. Place the slide flat on the rack provided at the lab reception window accompanied with a welllabelled request form.

3. Venepuncture for blood donation

Blood banks use various processes to try to prevent infections that can be transmitted by infected blood donation. One important measure to prevent infection is to recruit donors from populations that are known to have low rates of infection for bloodborne diseases, such as voluntary, unpaid donors and people with no history of intravenous drug use. A second measure is to ask donors a series of additional screening questions (these will vary by region) to help identify those who may be at higher risk of infection. Phlebotomists must adhere strictly to the rules for including and excluding blood donors. A third measure is to test donated blood for infections common in the area before processing it for use for various therapeutic purposes.

Blood donations should be collected only by trained and qualified blood transfusion services personnel.

Minimum requirements:

• Equipment:
  • All equipment used for collection of blood donations should be regularly calibrated, maintained and serviced, as required. Such equipment includes blood pressure monitors, scales, donor couches or chairs, blood collection monitors or mixers, blood bag tube sealers, blood transportation boxes and blood bank refrigerators.
  • Furniture and equipment in the area of blood donation and processing should be made of cleanable surfaces (e.g. vinyl rather than fabric). Containers used to transport supplies and specimens should also be cleanable by disinfectants such as sodium hypochlorite bleach solutions. Fabric or textile carriers should be machine washable.
• Location:
  • Premises should be of sufficient size for efficient operations, with separate areas for clean and dirty processes, clean running water, and surfaces cleanable by disinfectants.
  • Floors should not be carpeted.
  • Waiting areas should be outside the collection area, to minimize the risk of respiratory pathogens for workers.
  • All fixed and mobile blood donation sites should be safe, clean, hygienic and tidy, and should meet defined standards of environmental safety.
  • The donation sites should be organized in a way that ensures the safety of blood donors, staff and donated blood units, and avoids errors in the blood donation process.

Before a blood donation:

Blood donation should be voluntary; it should not involve duress, coercion or remuneration. Also, potential blood donors should be selected carefully, according to the national criteria for donor selection.

• The potential donor should be given pre-donation information, advice and counselling about the process of blood donation;
• A relevant history of the donor should be taken, covering health and high-risk behaviour,and including
  • History of mastectomy (blood should be taken from the arm opposite the site of surgery)
  • Current and recent medications or chronic infections;
  • History of prolonged bleeding or a past diagnosis of bleeding disorders;
  • History of previous donations, to ensure the waiting period is respected;
• A preliminary physical check-up of the donor should be done, including weight, blood pressure, signs of infection or scarring at potential sites
• The donor should be offered fluids, to help reduce the risk of fainting after blood donation
• The person should provide informed written consent, based on the national requirements.

Collecting blood:

For collection of blood for donation, use the procedure detailed in Chapter 2 for blood sampling (e.g. for hand hygiene and glove use), as far as it is relevant, and follow the six steps given below.

Identify donor and label blood collection bag and test tubes

• Ask the donor to state their full name.
• Ensure that:
  • the blood collection bag is of the correct type;
  • the labels on the blood collection bag and all its satellite bags, sample tubes and donor records have the correct patient name and number;
  • the information on the labels matches with the donor’s information.

Select the vein

• Select a large, firm vein, preferably in the antecubital fossa, from an area free from skin lesions or scars.
• Apply a tourniquet or blood pressure cuff inflated to 40–60 mm Hg, to make the vein more prominent.
• Ask the donor to open and close the hand a few times.
• Once the vein is selected, release the pressure device or tourniquet before the skin site is prepared.

Disinfect the skin:

Perform the venepuncture:

Perform venepuncture using a smooth, clean entry with the needle

• In general, use a 16-gauge needle, which is usually attached to the blood collection bag. Use of a retractable needle or safety needle with a needle cover is preferred if available, but all should be cut off at the end of the procedure rather than recapped.
• Ask the donor to open and close the fist slowly every 10–12 seconds during collection.
• Remove the tourniquet when the blood flow is established or after 2 minutes, whichever comes first.
• Closely monitor the donor and the injection site throughout the donation process – look for:
  • sweating, palor or complaints of feeling faint that may precede fainting;
  • development of a haematoma at the injection site;
  • changes in blood flow that may indicate the needle has moved in the vein, and needs to be repositioned.
• About every 30 seconds during the donation, mix the collected blood gently with the anticoagulant, either manually or by continuous mechanical mixing.
• Cut off the needle using a sterile pair of scissors.
• Collect blood samples for laboratory testing.

After a blood donation
Donor care
After the blood has been collected:
• ask the donor to remain in the chair and relax for a few minutes;
• inspect the venepuncture site; if it is not bleeding, apply a bandage to the site; if it is bleeding, apply further pressure;
• ask the donor to sit up slowly and ask how the person is feeling;
• before the donor leaves the donation room, ensure that the person can stand up without dizziness and without a drop in blood pressure;
• offer the donor some refreshments.
Blood unit and samples
• Transfer the blood unit to a proper storage container according to the blood centre requirements and the product.
• Ensure that collected blood samples are stored and delivered to the laboratory with.completed documentation, at the recommended temperature, and in a leak-proof, closed container

4. Arterial blood sampling:

For sampling from the radial artery using a needle and syringe, follow the steps outlined below.

1. Approach the patient, introduce yourself and ask the patient to state their full name.
2. Place the patient on their back, lying flat. Ask the nurse for assistance if the patient’s position needs to be altered to make them more comfortable. If the patient is clenching their fist, holding their breath or crying, this can change breathing and thus alter the test result.
3. Locate the radial artery by performing an Allen test for collateral circulation. If the initial test fails to locate the radial artery, repeat the test on the other hand. Once a site is identified, note anatomic landmarks to be able to find the site again. If it will be necessary to palpate the site again, put on sterile gloves.
4. Perform hand hygiene, clear off a bedside work area and prepare supplies. Put on an impervious gown or apron, and face protection, if exposure to blood is anticipated.
5. Disinfect the sampling site on the patient with 70% alcohol and allow it to dry.
6. If the needle and syringe are not preassembled, assemble the needle and heparinized syringe and pull the syringe plunger to the required fill level recommended by the local laboratory.
7. Holding the syringe and needle like a dart, use the index finger to locate the pulse again, inform the patient that the skin is about to be pierced then insert the needle at a 45 degree angle, approximately 1 cm distal to (i.e. away from) the index finger, to avoid contaminating the area where the needle enters the skin.
8. Advance the needle into the radial artery until a blood flashback appears, then allow the syringe to fill to the appropriate level. DO NOT pull back the syringe plunger.
9. Withdraw the needle and syringe; place a clean, dry piece of gauze or cotton wool over the site and have the patient or an assistant apply firm pressure for sufficient time to stop the bleeding. Check whether bleeding has stopped after 2–3 minutes. Five minutes or more may be needed for patients who have high blood pressure or a bleeding disorder, or are taking anticoagulants.
10. Activate the mechanisms of a safety needle to cover the needle before placing it in the ice cup. In the absence of a safety-engineered device, use a one-hand scoop technique to recap the needle after removal.
11. Expel air bubbles, cap the syringe and roll the specimen between the hands to gently mix it. Cap the syringe to prevent contact between the arterial blood sample and the air, and to prevent leaking during transport to the laboratory.
12. Label the sample syringe.
13. Dispose appropriately of all used material and personal protective equipment.
14. Remove gloves and wash hands thoroughly with soap and water, then dry using single-use towels; alternatively, use alcohol rub solution.
15. Check the patient site for bleeding (if necessary, apply additional pressure) and thank the patient.
16. Transport the sample immediately to the laboratory, following laboratory handling procedures.

Recommended order of draw for plastic vacuum tubes

Order of use	Type of tube/usual colour	Additive	Mode of action	Uses
1	Blood culture bottle (yellow-black striped tubes)	Broth mixture	Preserves viability of microorganisms	Microbiology – aerobes, anaerobes, fungi
2	Non-additive tube
3	Coagulation tubed (light blue top)	Sodium citrate	Forms calcium salts to remove calcium	Coagulation tests (protime and prothrombin time), requires full draw
4	Clot activator (red top)	Clot activator	Blood clots, and the serum is separated by centrifugation	Chemistries, immunology and serology, blood bank (cross-match)
5	Serum separator tube (red-grey tiger top or gold)	None	Contains a gel at the bottom to separate blood from serum on centrifugation	Chemistries, immunology and serology
6	Sodium heparin (dark green top)	Sodium heparin or lithium heparin	Inactivates thrombin and thromboplastin	For lithium level use sodium heparin, for ammonia level use either
7	PST (light green top)	Lithium heparin anticoagulant and a gel separator	Anticoagulants with lithium, separates plasma with PST gel at bottom of tube	Chemistries
8	EDTA (purple top)	EDTA	Forms calcium salts to remove calcium	Haematology, Blood Bank (cross-match) requires full draw
9	Blood tube (pale yellow top)	Acid-citrate-dextrose (ACD, ACDA or ACDB)	Complement inactivation	HLA tissue typing, paternity testing, DNA studies
10	Oxalate/fluoride (light grey top)	Sodium fluoride and potassium oxalate	Antiglycolytic agent preserves glucose up to five days	Glucoses, requires full draw (may cause haemolysis if short draw)

Rejection of samples

• Mislabelled/Unlabelled specimens
• Improper container
• Quantity not sufficient for testing
• Without test request
• Haemolysed
• Clotted (Where not indicated)

Techniques to Prevent Hemolysis:

• Mix all tubes with anticoagulant additives gently (vigorous shaking can cause hemolysis) 5-10 times.
• Avoid drawing blood from a hematoma; select another draw site.
• If using a needle and syringe, avoid drawing the plunger back too forcefully.
• Make sure the venipuncture site is dry before proceeding with draw.
• Avoid a probing, traumatic venipuncture.
• Avoid prolonged tourniquet application (no more than 2 minutes; less than 1 minute is optimal).
• Avoid massaging, squeezing, or probing a site.
• Avoid excessive fist clenching.
• If blood flow into tube slows, adjust needle position to remain in the center of the lumen.

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