The Critical Role of Additives in Blood Collection
Tube additives are substances placed inside evacuated blood collection tubes to preserve blood specimens for laboratory analysis. They ensure the stability of analytes, prevent clotting (or promote it for serum separation), and maintain cellular integrity from the point of collection to processing.[1] For the phlebotomy technician, understanding additives is critical because using the wrong tube or misidentifying its properties is a leading cause of specimen rejection and erroneous test results.
On the Phlebotomy Technician Certification exam, you are expected to know the color-coded system, the specific additive in each tube, the mechanism of action, and the order of draw required to avoid cross-contamination. This section breaks down that essential knowledge.
Key Concepts & Definitions
Understanding the function of each type of additive is the foundation of this topic. Below are the major categories used in clinical practice, as defined by the Clinical and Laboratory Standards Institute (CLSI).[1][3]
- Anticoagulants: Substances that prevent blood from clotting by binding calcium or inhibiting thrombin.
- EDTA (Ethylenediaminetetraacetic acid): Binds calcium. Preferred for hematology (CBC).
- Sodium Citrate: Binds calcium. Used for coagulation studies (PT/PTT). Maintains a specific 9:1 blood-to-additive ratio.
- Heparin (Lithium or Sodium): Inhibits thrombin. Used for plasma chemistry tests.
- Clot Activators: Substances that promote coagulation by activating platelets and the clotting cascade (e.g., silica, thrombin, glass particles). Used in serum (red/gold) tubes to speed up clotting time.
- Separation Gels: Synthetic, inert materials that form a physical barrier between cells and serum/plasma after centrifugation. Found in SST (Gold) and PST (Green/Gray) tubes.
- Preservatives: Chemicals that prevent the breakdown of specific analytes.
- Sodium Fluoride: A weak anticoagulant but a potent glycolytic inhibitor (preserves glucose). Found in Gray tubes.
- ACD (Acid Citrate Dextrose): Preserves red blood cells for blood banking and DNA testing.
Core Principles: The Color-Coded System & Order of Draw
The color of the tube stopper directly indicates the additive inside. Memorizing this system is non-negotiable for exam success and clinical safety. The Order of Draw is a standardized sequence established by the CLSI to prevent additive carryover from one tube to the next.[1][2]
High-Yield Tube Guide
| Stopper Color | Additive | Mechanism / Action | Primary Laboratory Use |
|---|---|---|---|
| Light Blue | Sodium Citrate (3.2%) | Binds calcium (anticoagulant) | Coagulation (PT/PTT/INR) |
| Red | None (or Silica) | Clot activator (glass/silica) | Serum chemistry, Serology |
| Gold / SST | Clot activator + Gel | Activates clotting; gel separates serum | Serum chemistry (most common) |
| Green | Sodium or Lithium Heparin | Inhibits thrombin (anticoagulant) | Plasma chemistry, STAT chemistry |
| Lavender / Pink | EDTA (K2 or K3) | Binds calcium (anticoagulant) | Hematology (CBC), Blood Bank (Pink) |
| Gray | Potassium Oxalate / Na Fluoride | Anticoagulant + Glycolytic inhibitor | Glucose, Lactic Acid, Alcohol |
| Yellow | ACD (Acid Citrate Dextrose) | Preserves RBCs / DNA | Blood banking, DNA studies |
The Order of Draw (CLSI GP41 Standard)[1]
Always follow this exact sequence during venipuncture to prevent interference. Failure to do so is a common cause of erroneous lab results.
- Blood Culture Tubes (SPS / Yellow) – Sterile, no additive.
- Light Blue Top (Coagulation tubes) – Additive carryover from other tubes can falsely activate clotting.
- Serum Tubes (Red, Gold/SST) – No additive or clot activator.
- Heparin Tubes (Green / PST) – Heparin can interfere with coagulation and chemistry tests if carried over.
- EDTA Tubes (Lavender / Pink) – EDTA carryover can chelate calcium falsely for other tests.
- Glycolytic Inhibitor Tubes (Gray) – Fluoride/oxalate carryover is highly disruptive.
Safety Precautions & Common Complications
Additives are powerful reagents. Failure to manage them correctly poses risks to both the patient and the integrity of the lab results.[4][5]
- Underfilling Tubes: Most critical for Light Blue (Citrate) tubes. An incorrect blood-to-additive ratio (less than 9:1) leads to excess citrate, which binds more calcium, falsely prolonging PT/PTT results. The tube must be filled completely to the fill line.[2]
- Inadequate Mixing: Inverting tubes gently but thoroughly 3-10 times (depending on the manufacturer) is essential. Failure to mix allows visible clotting or microclots to form, ruining the sample for hematology and coagulation analyzers.
- Additive Carryover (Order of Draw Errors): As noted above, drawing a Lavender (EDTA) tube before a Light Blue (Citrate) tube introduces EDTA into the coagulation sample, falsely lowering calcium and inhibiting clotting factors.
- Failing to Invert Gel Tubes: Gel SST/PST tubes require thorough inversion to ensure the clot activator and gel are mixed with the blood. If not mixed, the gel may fail to form a proper barrier during centrifugation, leading to cell/serum interface instability.
Exam Tips & High-Yield Points
Certification exams heavily test tube identification, additive mechanisms, and the order of draw. Use these memory aids to lock in the information.
Memory Aids
- "Lavender for Live Cells": Lavender tube (EDTA) prevents hemolysis and preserves cell morphology for the CBC.
- "Blue before you go, Green before the sheen, Lavender after the rest": A quick rhyme for the first three steps of the order of draw (Blue, Red/Gold, Green).
- "Fluoride = Fasting Glucose": Gray tube (Sodium Fluoride) is the go-to for accurate glucose levels because it halts glycolysis.
- "Coagulation = Citrate": Both start with "C". The Light Blue tube is for coagulation studies (PT/PTT).
Common Exam Questions Focus On:
- Identifying the additive: "Which additive is in a light blue top tube?" Answer: Sodium Citrate.
- The consequence of a wrong tube: "What happens if you draw a green top tube for a PT/PTT?" Answer: Heparin in the tube will inhibit thrombin, causing a falsely prolonged result.
- The reason for the order of draw: "Why is a light blue top tube drawn BEFORE a green top tube?" Answer: To avoid heparin carryover contamination into the coagulation sample.
- Additive Function: "What is the primary function of EDTA?" Answer: Binds calcium to prevent coagulation.
References & Sources
The clinical standards and academic guidelines used to compile this study guide section are listed below.
- Clinical and Laboratory Standards Institute (CLSI). GP41: Collection of Diagnostic Venous Blood Specimens. 7th ed. CLSI; 2017. https://en.wikipedia.org/wiki/Clinical_and_Laboratory_Standards_Institute
- Clinical and Laboratory Standards Institute (CLSI). GP42: Collection, Transport, and Processing of Blood Specimens for Testing Plasma-Based Coagulation Assays. 6th ed. CLSI; 2008. https://clsi.org/standards
- McCall RE, Tankersley CM. Phlebotomy Essentials. 7th ed. Wolters Kluwer; 2020. https://samples.jblearning.com/9781284209945/9781284209501_FMxx_i_xxxiv_Secured.pdf
- Ernst DJ, Ernst C. Phlebotomy Exam Review. 3rd ed. Skyward Publishing; 2019. https://repository.stikesbcm.ac.id/id/eprint/177/1/Phlebotomy.pdf
- Garza D, Becan-McBride K. Phlebotomy Handbook: Blood Collection Essentials. 10th ed. Pearson; 2018. https://www.r2library.com/