Blood Sample Collection

Table of contents

Introduction

Blood sample collection is a vital procedure in laboratory medicine for diagnosing, monitoring, and predicting diseases.
The accuracy of laboratory results depends greatly on the quality of the collected blood specimen.
It is performed by trained healthcare professionals who follow standardized techniques.
Aseptic methods are used during the procedure to prevent contamination and infection.
Proper patient preparation, including identification and checking fasting status, is essential.
Blood collection forms a major part of the pre-analytical phase where most errors can occur, making correct technique crucial.

Types of Blood Sample Collection

a) Venous Blood Collection

Most commonly used method
Collected from the median cubital, cephalic, or basilic veins
Used for serum chemistry, hematology, immunology, microbiology, and specialized assays

b) Capillary Blood Collection

Taken from fingertip (adults) or heel (infants)
Useful in glucose testing, neonatal screening, POCT (Point-of-Care Testing)

c) Arterial Blood Collection

Performed for arterial blood gas (ABG) analysis
Requires expertise due to higher risk

Preparation for Blood Collection

Identify the Patient

Verify patient identity using at least two identifiers, such as:
- Full name
- Date of birth
- Patient identification number
Explain the procedure to the patient to obtain verbal consent and address any concerns they may have.

Patient Preparation

Proper preparation ensures accurate and reproducible results:

Confirm patient identity (two-step verification)
Explain the procedure to reduce anxiety
Check for fasting requirements
Assess hydration status
Avoid sample collection from IV infusion arm
Ensure patient is seated or lying down

Equipment for sample collection

Ensure all necessary materials are prepared before starting the procedure:

Vacutainer tubes (correct type based on the tests required)
Needles (typically 21 or 22 gauge for adults)
Tourniquet
Alcohol swab or antiseptic
Sterile gauze or cotton
Adhesive bandage
Gloves
Sharps container for needle disposal
Labels for the tubes (with patient information)

Choose the Collection Site

Skin puncture

Skin puncture, or capillary blood collection, is a common technique to obtain small amounts of blood for various tests. This method is often preferred for infants, children, or individuals with limited access to veins.

Steps Involved in Skin Puncture:

Preparation:
- Gather necessary materials: sterile lancets, alcohol swabs, cotton balls, bandages, and appropriate collection tubes or devices.
- Ensure the patient is comfortable and informed about the procedure.
Site Selection:
- Choose a suitable site, typically the fingertip or heel, for infants.
- Clean the area with alcohol and allow it to dry completely.
Puncture:
- Use a sterile lancet to make a shallow puncture. Avoid going too deep to prevent tissue damage.
- Gently squeeze the area to encourage blood flow.
Blood Collection:
- Place the collection device against the puncture site to collect the blood.
- Follow any specific instructions provided by the laboratory or test kit.
Site Care:
- Apply pressure to the puncture site with a cotton ball until the bleeding stops.
- Cover the site with a bandage if necessary.

Precautions and Considerations:

Sterility: Maintain a sterile environment to prevent infection.
Depth: Avoid puncturing too deeply, which can cause pain and tissue damage.
Pressure: Excessive pressure can hemolyze (rupture) red blood cells, affecting test results.
Volume: Ensure sufficient blood is collected for the required tests.
Consent: Obtain consent from the patient or guardian, especially for children.

Vein puncture

Common Sites:

- The antecubital fossa (inner elbow) is the most common area, with veins like the median cubital, cephalic, and basilic veins being preferred.
- In some cases, veins in the dorsal hand or wrist may be used if the arm veins are difficult to access.

Steps for Venipuncture

Position the Patient

Have the patient sit or lie down with their arm extended on a flat surface.
Ensure the patient’s arm is relaxed and supported, with the palm facing upward.

Apply the Tourniquet

Apply the tourniquet 3-4 inches above the puncture site to engorge the veins.
The tourniquet should not be left on for more than 1 minute to avoid hemoconcentration, which can affect test results.

Select the Vein

Palpate the vein using your fingertips to feel for a firm, round, and elastic vein.
Veins are typically easier to locate in well-hydrated patients.

Clean the Site

Clean the selected puncture site with an alcohol swab in a circular motion, starting from the center and working outward.
Allow the area to air-dry to avoid introducing alcohol into the sample.

Insert the Needle

Put on gloves to maintain an aseptic technique.
Uncap the needle and hold it at a 15-30° angle with the bevel facing up.
Insert the needle into the vein with a smooth, quick motion. The needle should enter with a slight downward angle.

Collect the Sample

Attach the appropriate vacutainer tube to the needle holder to collect blood.
- Order of Draw: Blood should be drawn in a specific sequence to avoid contamination between additives in different tubes. The typical order of draw is:

Blood culture tubes (yellow or special bottles)
Citrate tubes (light blue top)
Serum tubes (red or gold top)
Heparin tubes (green top)
EDTA tubes (lavender top)
Fluoride/oxalate tubes (gray top)

Gently invert the tubes 5-10 times after collection to mix the blood with the anticoagulant or preservative.
- Avoid shaking to prevent hemolysis.

Release the Tourniquet

Release the tourniquet once blood flow is established to prevent prolonged pressure on the vein, which can cause hemolysis or tissue damage.

Remove the Needle

After collecting the required blood volume, carefully remove the needle while applying light pressure to the site with sterile gauze or cotton.
Dispose of the needle immediately into a sharp container.

Apply Pressure and Bandage

Apply firm pressure to the puncture site using sterile gauze for 1-2 minutes to stop bleeding.
Once the bleeding has stopped, apply an adhesive bandage.

Post-Collection Procedure

Label the Tubes

Immediately label each collection tube with the patient’s:
- Full name
- Date of birth
- Time and date of collection
- Test(s) requested
Ensure the information on the tubes matches the patient’s identification.

Transport and Storage

Store and transport the samples according to the specific requirements of the tests:
- Room temperature for most haematological and coagulation studies.
- Refrigeration at 4°C for some samples (e.g., blood gases or specific chemistry tests).
- Avoid exposing samples to excessive heat or freezing, which can affect the integrity of certain blood components.

Documentation

Document the procedure in the patient’s medical record, including:
- The date and time of collection
- The collection site
- The type of test ordered
- Any issues encountered during the collection process

Common Errors and Precautions

Hemolysis

Cause: Vigorous shaking of tubes, using a small-gauge needle, or prolonged tourniquet application.
Effect: This can lead to inaccurate test results like potassium levels, LDH, and haemoglobin.

Contamination

Cause: Not following the proper drawing order, reusing needles, or improper cleaning of the puncture site.
Effect: Can introduce additives from one tube into another, leading to inaccurate results (e.g., false elevated potassium in a serum sample).

Clotting in Anticoagulated Samples

Cause: Failure to invert tubes with anticoagulant properly.
Effect: Leads to clotting of the sample, rendering it unsuitable for haematological testing.

Fainting (Syncope)

Precaution: Always monitor the patient for signs of fainting or dizziness during and after the procedure.
Have the patient seated or lying down during collection to prevent falls.

Collection & Preservation for Specific Hematological Investigations

Complete Blood Count (CBC)

Test Description: Measures various blood components, including RBC count, WBC count, haemoglobin, hematocrit, and platelet count.

Tube Type: EDTA (Lavender Top)
- Anticoagulant: Ethylenediaminetetraacetic acid (EDTA).
- Mode of Action: EDTA chelates calcium, preventing clotting and preserving cellular morphology.
Sample Handling:
- Ensure gentle mixing of the blood with the anticoagulant by inverting the tube 8-10 times immediately after collection.
- Avoid vigorous shaking, which can cause hemolysis (rupture of red blood cells).
Storage:
- Room temperature: Stable for up to 6 hours for routine CBC tests.
- Refrigeration (4°C): Can preserve samples for up to 24 hours for most parameters but may affect some cell counts like platelets.

Coagulation Studies (e.g., PT, aPTT)

Test Description: Assesses blood clotting times to detect deficiencies in the clotting factors or monitor anticoagulant therapy (e.g., warfarin, heparin).

Tube Type: Sodium Citrate (Light Blue Top)
- Anticoagulant: 3.2% Sodium Citrate.
- Mode of Action: Binds calcium reversibly, preventing coagulation.
Sample Handling:
- The blood-to-anticoagulant ratio must be 9:1 (blood) to ensure accurate test results.

- Fill the tube to the marked level to maintain the correct anticoagulant ratio.
Storage:
- Room temperature: Analyze samples within 2-4 hours for accurate coagulation results.
- Refrigeration: Not recommended, as it can activate clotting factors and skew results.

Erythrocyte Sedimentation Rate (ESR)

Test Description: Measures the rate at which red blood cells settle in a column of blood, which can indicate inflammation or infection.

Tube Type: Sodium Citrate (Black Top) or EDTA (Lavender Top).
- Anticoagulant: Sodium Citrate (for ESR tubes) or EDTA.
Sample Handling:
- Gently invert the tube several times to ensure proper mixing of blood and anticoagulant.
Storage:
- Room temperature: Analyze within 2 hours.
- Refrigeration (4°C): Samples can be stored for up to 24 hours, but allow the sample to return to room temperature before analysis.

Blood Smear for Peripheral Blood Film

Test Description: Used to examine blood cell morphology and detect abnormalities in RBCs, WBCs, and platelets.

Tube Type: EDTA (Lavender Top).
Sample Handling:
- Prepare the smear within 2-3 hours of collection to avoid changes in cell morphology (e.g., red cell crenation or WBC degradation).
- Smears should be air-dried quickly and fixed with methanol for staining.
Storage:
- Room temperature: Prepare smears immediately; store the slides at room temperature once prepared.

Reticulocyte Count

Test Description: Measures immature red blood cells (reticulocytes) to assess bone marrow function and the body’s response to anaemia.

Tube Type: EDTA (Lavender Top).
Sample Handling:
- Gently invert the tube to ensure proper mixing with the anticoagulant.
- Process the sample within 4-6 hours for best results.
Storage:
- Room temperature: Stable for up to 6 hours.
- Refrigeration (4°C): Samples can be stored for up to 24 hours.

Bone Marrow Aspiration

Test Description: Used to diagnose hematologic disorders like leukaemia, anaemia, and myelodysplastic syndromes.

Tube Type: EDTA (Lavender Top) or Heparin (Green Top) for certain cytogenetic tests.
Sample Handling:
- Bone marrow aspirates should be processed immediately for smear preparation.
- If cytogenetic analysis is required, collect in heparinized tubes to preserve cell viability.
Storage:
- Room temperature: Smears should be prepared as soon as possible after collection.

Sample Preservation and Storage Conditions

Refrigeration (4°C):

Used for short-term preservation of most haematology samples (except coagulation tests).
Extends the stability of cell morphology and certain analytes, especially for tests like CBC and reticulocyte count.

Freezing:

Not recommended for whole blood samples, as it can lead to hemolysis and cellular damage.
Some components, like plasma for coagulation studies, may be frozen if separated from cells.

Room Temperature:

For tests like coagulation studies, ESR, and blood smears, samples should generally be processed within 2-4 hours of collection.

Avoiding Hemolysis:

Hemolysis can occur due to improper handling (e.g., vigorous shaking, small-gauge needle use, or excessive force during venipuncture).
Hemolysis can interfere with tests, leading to inaccurate results, particularly in tests like potassium levels, LDH, and haemoglobin.

MCQs

Which vein is most commonly used for venipuncture?
A. Basilic vein
B. Cephalic vein
C. Median cubital vein
D. Femoral vein
What is the first step before collecting any blood sample?
A. Apply tourniquet
B. Clean the site
C. Identify the patient
D. Insert the needle
Which color tube is used for CBC testing?
A. Red
B. Lavender
C. Blue
D. Yellow
The lavender-top tube contains:
A. Heparin
B. EDTA
C. Sodium fluoride
D. Citrate
Which tube is used for coagulation studies (PT, APTT)?
A. Red
B. Blue
C. Green
D. Grey
Which anticoagulant prevents clotting by binding calcium?
A. EDTA
B. Heparin
C. Thrombin
D. Polymer gel
Capillary blood is usually collected from which site in adults?
A. Toe
B. Ear lobe
C. Finger
D. Heel
Capillary blood in infants is collected from:
A. Finger
B. Heel
C. Wrist
D. Forearm
What is the ideal angle for inserting a needle in venipuncture?
A. 15–30°
B. 45–60°
C. 75–90°
D. 5–10°
The order of draw begins with:
A. EDTA tubes
B. Coagulation tubes
C. Blood culture bottles
D. Serum tubes
Which factor can cause hemolysis during collection?
A. Using a large-bore needle
B. Gentle mixing
C. Vigorous shaking of tube
D. Correct storage
Tourniquet should not be applied for more than:
A. 10 seconds
B. 20 seconds
C. 60 seconds
D. 5 minutes
Which needle size is commonly used in adult venipuncture?
A. 16G
B. 18G
C. 21–23G
D. 30G
Which of the following may lead to a falsely elevated potassium level?
A. Hemolysis
B. Using EDTA tube
C. Delayed centrifugation
D. All of the above
The grey-top tube contains:
A. Fluoride + oxalate
B. EDTA
C. Heparin
D. No additive
Which sample is required for blood glucose estimation?
A. Serum
B. EDTA blood
C. Fluoride plasma
D. Citrated blood
The process of removing serum from a clotted sample is called:
A. Coagulation
B. Plasma separation
C. Serum separation
D. Decanting
Syncope refers to:
A. Bleeding
B. Fainting
C. Infection
D. Hemolysis
Which antiseptic is commonly used before blood collection?
A. Phenol
B. Spirit + iodine
C. Turpentine
D. Silver nitrate
For blood culture, the skin must be cleaned with:
A. Soap
B. 70% alcohol only
C. Chlorhexidine
D. Normal saline
Which sample is preferred for ESR (Westergren method)?
A. Heparinized blood
B. Citrated blood
C. Serum
D. Fluoride blood
Hemoconcentration is caused by:
A. Prolonged tourniquet application
B. Short tourniquet application
C. Using small needle
D. Low temperature
How many times should an EDTA tube be inverted after collection?
A. 1–2 times
B. 8–10 times
C. 20 times
D. Shake vigorously
Which is the safest method to dispose of used needles?
A. Break manually
B. Recap the needle
C. Discard in puncture-proof container
D. Throw in dustbin
Which tube gives a serum sample?
A. Red-top
B. Lavender
C. Green
D. Grey
Plasma differs from serum because it contains:
A. Red cells
B. Fibrinogen
C. Platelets
D. Water
Which factor increases the risk of hemolysis?
A. Cold storage
B. High pressure during blood withdrawal
C. Using vacuum tubes
D. Correct needle angle
Which specimen is required for arterial blood gas (ABG)?
A. Venous blood
B. Capillary blood
C. Arterial blood
D. Serum
The most common artery used for ABG collection:
A. Carotid artery
B. Ulnar artery
C. Radial artery
D. Femoral artery
Before radial artery puncture, which test is performed?
A. Coagulation test
B. Allen’s test
C. Hemoglobin test
D. ESR test
Which additive prevents clotting by inhibiting thrombin?
A. EDTA
B. Citrate
C. Heparin
D. Fluoride
Ideal position for the patient during venipuncture:
A. Standing
B. Lying or sitting comfortably
C. Running
D. Leaning forward
What should be done if a hematoma develops during collection?
A. Continue collection
B. Apply pressure and stop
C. Insert needle deeper
D. Change nothing
Which factor affects blood test results?
A. Fasting
B. Exercise
C. Stress
D. All of the above
Which tube should be filled completely to maintain correct ratio?
A. EDTA
B. Citrate (blue-top)
C. Fluoride
D. Heparin

Answer Key