Overview
Blood collection is one of the most fundamental and frequently performed procedures in a hematology laboratory. The quality of every test result depends entirely on the quality of the specimen — and that begins at the moment of collection.
Blood can be collected as venous blood from the antecubital vein (preferred for most tests), or as capillary blood from the finger or heel (used when venipuncture fails or in infants). Each method has specific indications, techniques, and precautions.
Learning Objectives
- Describe the complete technique of venous blood collection using a syringe and needle
- Describe the technique of venous blood collection using the vacutainer system
- Identify the different types of vacuum tubes by cap colour and their anticoagulants
- Describe the method of collecting capillary blood from finger and heel
- Explain the three main anticoagulants used in the hematology laboratory and their mechanisms
- Differentiate between whole blood, serum and plasma and state their uses
Clinical Story
A doctor orders a coagulation test (PT/APTT) and a complete blood count for a post-operative patient. The phlebotomist collects all samples into purple EDTA tubes. The coagulation samples are rejected and the patient must be re-bled. A small knowledge gap — not knowing which tube to use — caused patient distress, delayed results, and wasted resources. This lesson prevents that mistake.
Collection of Venous Blood
Blood is withdrawn from the antecubital vein or any well-identified forearm vein. The selected vein must be large, readily accessible, and sufficiently close to the surface to be seen and palpated.
Site Preparation
Clean the skin around the identified vein with 70% isopropyl alcohol in a circular fashion, beginning at the puncture site and moving outward. Allow to dry spontaneously. Do not touch the venipuncture site after cleaning.
Apply a tourniquet 3–4 inches above the venipuncture site. Ask the patient to make a fist several times. Veins can also be made more prominent by allowing the arm to hang down for 2–3 minutes or gently slapping the site. Release the tourniquet as soon as blood enters the collection vessel.
Syringe and Needle Method
Wash hands thoroughly. Label the collection tube with the patient's name and hospital number before drawing blood. Attach needle to syringe; check it works smoothly. Keep needle capped until use.
With the needle bevel up and parallel to the skin surface, insert into the vein at a shallow angle. Appearance of blood in the needle hub confirms successful venous entry. Release the tourniquet immediately.
Withdraw the piston slowly to avoid frothing (haemolysis). Collect the required amount.
Place a sterile gauze pad over the puncture site. Withdraw the needle while simultaneously applying pressure. Apply light pressure until bleeding stops, then cover with an adhesive dressing.
Deliver blood gently into the specified tube. Cap firmly. Do NOT shake — invert gently to mix anticoagulant.
Destroy the needle immediately in a needle destroyer. NEVER recap, bend or break. Place used swab, syringe and contaminated material in a puncture-resistant container.
Vacutainer System
The Vacutainer system consists of a double-pointed needle, a plastic holder/adapter, and several vacuum tubes with colour-coded rubber stoppers. Blood flows directly from vein to tube under vacuum. No blood transfer is required, reducing contamination risk.
Label all tubes with patient identification. Place the vacuum tube in the holder until the stopper top is level with the guideline. Attach disposable needle.
Place the patient's arm in a downward position to reduce the risk of backflow of anticoagulant into the patient's circulation.
Insert needle into vein. Push tube into needle to puncture the stopper/vacuum seal. Blood flows in automatically. Release tourniquet as soon as blood appears.
If more samples are needed, remove the filled tube and insert the next. The needle remains in the vein. While successive tubes fill, invert previous tubes gently to mix — do NOT shake vigorously (causes haemolysis).
Remove holder, cover site with sterile swab, apply pressure until bleeding stops. Destroy needle in destroyer without recapping.
Backflow of anticoagulant from the tube into the patient's vein can cause an adverse reaction. Always keep the arm downward during vacutainer collection.
Types of Vacuum Tubes — Colour Code
| Cap Colour | Anticoagulant / Additive | Test Used For | Notes |
|---|---|---|---|
| Purple / Lavender | EDTA (dipotassium salt) | Complete Blood Count (CBC), reticulocyte count, peripheral blood film | Prevents clotting by chelating calcium |
| Red | None (plain tube) | Tests requiring serum (biochemistry, serology) | Blood clots; serum separated by centrifugation |
| Blue | Sodium Citrate | Coagulation tests (PT, APTT, fibrinogen) | Blood:anticoagulant ratio must be exactly 9:1 |
| Grey | Fluoride (+ oxalate) | Blood glucose, lactate | Fluoride inhibits glycolysis — preserves glucose levels |
"Purple for CBC, Blue for clotting, Red for serum, Grey for glucose." A simple sentence that covers the four tubes you use every day.
Collection of Capillary Blood
Capillary blood is obtained by skin puncture with a needle or lancet. It is used in small children (infants), very obese adults in whom venipuncture fails, and for point-of-care testing. Samples are suitable for peripheral blood films, haematocrit, and haemoglobin estimation.
| Patient | Collection Site |
|---|---|
| Adults | Lateral side of the tip of the 3rd or 4th finger |
| Infants | Deep puncture of the plantar surface of the heel |
Capillary Collection Procedure
Clean the area with 70% alcohol and allow to dry spontaneously.
Puncture the skin to a depth of 2–3 mm with a sterile disposable lancet or needle.
Wipe away the first drop of blood (it contains tissue fluid which dilutes the sample). Squeeze gently to allow free flow. In a good puncture, large drops should exude spontaneously. Do NOT squeeze firmly — this gives unreliable results due to dilution with tissue fluid.
Anticoagulants Used in Hematology
Primary use: Complete Blood Counts (CBC). The dipotassium (K₂) salt is preferred — it is more soluble and is used in solid form coated on the inside of the tube.
Mechanism: Chelates (removes) calcium ions from the blood. Calcium is essential for coagulation, so removing it prevents clotting.
Recommended concentration: 1.50 ± 0.25 mg/mL of blood (ICSH standard).
If EDTA is used in excess (too little blood added to the tube):
• Causes shrinkage of RBCs and leucocytes → falsely low haematocrit
• Significant decrease in haematocrit, increase in MCHC
• Platelets swell and disintegrate → falsely high platelet count
Always add the correct volume of blood to the EDTA tube.
Primary use: Coagulation studies (PT, APTT, fibrinogen).
Mechanism: Binds calcium, preventing coagulation.
Critical ratio for coagulation: 9 volumes blood : 1 volume anticoagulant (0.5 mL citrate + 4.5 mL blood). The ratio is critical — if incorrect, all coagulation results will be wrong.
Also used for ESR (Westergren method): 4 volumes blood : 1 volume sodium citrate.
The citrate:blood ratio directly affects the final calcium concentration in the test. An incorrect ratio means the test is performed with the wrong amount of anticoagulant — invalidating all clotting factor results. Always ensure the tube is filled to the correct level (the line marked on the tube).
Primary use: Osmotic fragility testing, red cell enzyme studies (e.g. G6PD), immunophenotyping.
Salt used: Sodium or lithium heparin at 10–20 IU/mL blood.
Advantage: Does not change red cell size — best anticoagulant for osmotic fragility.
✗ NOT suitable for CBC — causes platelet and leucocyte clumping → false counts
✗ NOT for peripheral blood films — gives a faint blue background after Romanowsky staining, interfering with cell morphology assessment
Types of Samples
| Sample Type | How Obtained | Contains | Used For |
|---|---|---|---|
| Whole Blood | Anticoagulated blood (EDTA) | All cells + plasma + clotting factors | CBC, reticulocyte count, peripheral blood film |
| Serum | Blood allowed to clot → centrifuge at 1200g / 10 min | Plasma minus clotting factors | Biochemical tests, serum protein electrophoresis |
| Plasma | Anticoagulated blood → centrifuge | Fluid portion with clotting factors intact | Coagulation studies (PT, APTT) |
Common Errors & How to Avoid Them
Cause: Withdrawing piston too fast, shaking tube vigorously, using too small a needle, contaminating with water or alcohol.
Effect: Red cells lyse → falsely high potassium, falsely low RBC count. Sample rejected.
Prevention: Withdraw piston slowly, invert gently, ensure adequate drying of site before collection.
Cause: Confusing tube colours, labelling error, using EDTA for coagulation tests.
Effect: Invalid results, patient re-bleed required.
Prevention: Always verify tube colour against the requested test. Check label before filling.
Cause: Underfilling or overfilling the citrate tube.
Effect: All coagulation results are invalid.
Prevention: Fill the tube exactly to the marked fill line. Never use a partially filled tube for coagulation testing.
Cause: Leaving tourniquet on too long during difficult venipuncture.
Effect: Haemoconcentration — falsely elevated RBC, Hb, haematocrit values.
Prevention: Release tourniquet as soon as blood enters the collection vessel.
Cause: Collecting the first drop of capillary blood.
Effect: First drop contains tissue fluid → dilutes the sample → falsely low results.
Prevention: Always wipe away the first drop. Collect only from freely flowing blood.
Laboratory Tips
Always verify the patient's identity before collection. Check that the name and details on the requisition form match the patient. Write or print the label before collection — never label after from memory.
For patients with difficult veins: ask them to make a fist repeatedly, let the arm hang down for 2–3 minutes to engorge veins with gravity, or apply a warm compress for 5 minutes to dilate superficial veins.
When collecting multiple tubes: Blood culture → Blue (citrate) → Red (plain) → Grey (fluoride) → Purple (EDTA). The order prevents cross-contamination of anticoagulants between tubes. "BC Before Red Grapes Purple" — a rough mnemonic to get you started.
Interactive Quiz
5 QuestionsFlashcards
Tap to flipClinical Case Study
James is day 3 post-surgery and on warfarin anticoagulation therapy. His surgeon requests a PT/INR and CBC. The phlebotomist collects 2 mL into a blue citrate tube and 3 mL into a purple EDTA tube. The coagulation laboratory rejects the citrate sample and reports it as unacceptable.
The citrate tube was underfilled — only 2 mL of blood instead of 4.5 mL. This means the blood:anticoagulant ratio is wrong (4:1 instead of 9:1). Excess citrate over-anticoagulates the sample, producing a falsely prolonged PT. The sample must be rejected and the patient re-bled with a correctly filled tube.
- →The blood:citrate ratio is non-negotiable — an incorrect ratio invalidates all coagulation results
- →Specimen rejection protects the patient from incorrect clinical decisions
- →Quality begins at the moment of collection — every tube must be filled correctly
Quick Revision
10-Minute ReviewKey Takeaways
- Blood is collected from the antecubital vein using a syringe or vacutainer system
- The skin must be cleaned with 70% alcohol; tourniquet applied 3–4 inches above the site
- Needles must never be recapped, bent or broken — always destroyed immediately
- Vacuum tube cap colour indicates the anticoagulant: Purple=EDTA, Blue=Citrate, Red=plain, Grey=Fluoride
- EDTA chelates calcium to prevent clotting; in excess it causes cell shrinkage and false platelet counts
- Citrate blood ratio for coagulation MUST be exactly 9:1
- Heparin must not be used for CBC or peripheral blood films
- Capillary blood: adults from 3rd/4th finger; infants from heel; always discard first drop
- Serum lacks clotting factors; plasma retains them — each has specific uses