Order of Draw & Tube Additives – Comprehensive Study Notes

Order of Draw – Core Principles

• Purpose: Maintain specimen integrity by preventing additive carry-over and ensuring accurate test results.
• Sequence is mandatory; deviating can lead to hemolysis, clot formation, or cross-contamination of additives.
• Mnemonic (one of the many in circulation): “Cultures Love Rich Students Giving Lavish Gifts Regularly Yearly.”
  • C = Cultures
  • L = Light Blue
  • R = Red
  • S = SST (Tiger/Gold)
  • G = Green
  • L = Lavender
  • G = Gray
  • R = Royal Blue
  • Y = Yellow

Tube-by-Tube Details

CULTURE (Yellow SPS)

• Additive: SPS (Sodium Polyanethol Sulfonate)
• Primary Purpose:
  • Blood cultures (bacterial, fungal)
  • DNA testing (when sterility is essential)
• Mode of Action: SPS inactivates complement & certain antibiotics; promotes organism recovery.
• Special Handling:
  • Strict aseptic collection; often collected with aerobic & anaerobic bottles.
  • Requires thorough site disinfection (chlorhexidine or povidone-iodine).

LIGHT BLUE

• Additive: Sodium Citrate (usually $3.2\%$ or $3.8\%$)
• Tube Fill: Must be filled to indicated volume (9:1 blood-to-anticoagulant ratio) to ensure valid coag studies.
• Common Tests:
  • Coagulation studies overall
  • $PT/INR$ (Prothrombin Time / International Normalized Ratio)
  • $APTT$ or $PTT$
  • D-Dimer
  • Fibrinogen assays

RED (Glass or Plastic “Plain” Tube)

• Additive: None (glass) or clot activator (plastic) but still classified as additive-free for order purposes.
• Serum is required when additives might interfere.
• Typical Uses:
  • Serum determinations for infectious disease titers
  • STD panels
  • Drug levels when gel separator is contraindicated

SST – TIGER / GOLD (Serum Separator Tube)

• Additive: Thixotropic Gel plus clot activator
• Function: Upon centrifugation gel forms an inert barrier between serum & cells.
• Key Test Families:
  • Comprehensive metabolic panels (CMP)
  • Basic metabolic panels (BMP) & electrolyte panels
  • Lipid profiles (chol, HDL, LDL, triglycerides)
  • Endocrine markers (TSH, Vitamin $D$, Vitamin $B_{12}$, PSA, etc.)
  • "Panels, Lipids, TSH, CMP, PSA" explicitly listed in transcript
• Advantages: Eliminates need to pour-off serum; minimizes cellular metabolism of analytes.

GREEN

• Additive Choices: Sodium Heparin or Lithium Heparin
• Anticoagulant Mechanism: Heparin potentiates antithrombin III → inhibits thrombin & Factor Xa.
• Recommended Analyses:
  • STAT chemistry (electrolytes) because plasma can be tested immediately
  • Platelet counts & some specialized coag tests
  • Mentioned in transcript: “Electrolytes, Coagulation Studies, Platelet Count”

LAVENDER (a.k.a. Purple)

• Additive: EDTA (Ethylenediaminetetraacetic acid) – chelates $Ca^{2+}$ preventing clotting.
• Signature Tests:
  • Complete Blood Count (CBC)
  • White Blood Cell (WBC) differential
  • Erythrocyte Sedimentation Rate (ESR / Sed Rate)
  • Hemoglobin $A_{1c}$
  • Other whole-blood hematology assays
• Note: Inversion $8$× post-collection to ensure proper mixing (standard hematology practice).

GREY

• Additives:
  • Sodium Fluoride (glycolysis inhibitor)
  • Potassium Oxalate (anticoagulant)
• Major Uses:
  • Blood glucose level determinations (prevents in-vitro glycolysis)
  • GGT (Gamma-Glutamyl Transferase)
  • ETOH (blood alcohol) testing – fluoride stabilizes alcohol concentration
• Pre-analytic Caution: For legal ETOH draw, avoid alcohol swabs; use iodine or soap.

ROYAL BLUE

• Additive Variants:
  • Sodium Heparin (for plasma trace-element studies)
  • Sodium EDTA (for whole-blood trace metals)
• Specialty Niche: Toxicology & trace-element testing where ultra-low contamination is critical
• Example Analytes: Lead ($Pb$), Zinc ($Zn$), Copper ($Cu$), drug levels such as Dilantin (phenytoin) & Digoxin noted in transcript.
• Distinct Manufacturing: Silica-washed glass & rubber to minimize metal contamination.

YELLOW (ACD)

• Additive: Acid Citrate Dextrose (ACD)
  • Citrate = anticoagulant; dextrose = preserves cell viability.
• Core Tests:
  • Paternity testing (HLA typing)
  • ABO/Rh blood typing & extended immunohematology profiles
• Storage: Designed to maintain cellular integrity for DNA or antigen studies up to $21$ days under refrigeration.

Practical / Procedural Connections

• Order of draw mirrors CLSI standards to avoid additive carry-over:
  1. Blood Cultures/Yellow SPS
  2. Light Blue (Citrate)
  3. Red (No additive / glass)
  4. SST (Tiger/Gold)
  5. Green (Heparin)
  6. Lavender (EDTA)
  7. Grey (Fluoride/Oxalate)
  8. Royal Blue (Heparin/EDTA) – drawn after tubes with similar additives but before ACD in some labs
  9. Yellow ACD (special genetic use)
• Always verify local policy—minor variations exist, especially when specialized tubes (e.g., pink EDTA for blood bank) are introduced.
• Mix gently: Standard $4$–$8$ inversions—vigorous shaking can hemolyze cells; insufficient mixing can clot samples.

Ethical & Quality Considerations

• Patient Identification: Use two identifiers (full name, DOB or MRN) before any draw to comply with CAP/JCAHO.
• Chain of Custody: Mandatory for ETOH or forensic toxicology.
• Pre-Analytic Error Mitigation:
  • Correct order of draw reduces $\approx 30\%$ of sample rejections linked to additive contamination (literature estimate).
  • Proper tube volume & mixing directly influence test validity.
• Infection Control: Blood culture collection demands max sterility to avoid false positives → reduces unnecessary antibiotic use.

Quick Comparison Chart (from Transcript)

• Culture / SPS → Blood Cultures, DNA
• Light Blue / Sodium Citrate → Coag (PT/INR, D-Dimer)
• Red / None → Serum determinations, STDs, Titers, Vitamin $D/B_{12}$
• SST (Tiger/Gold) / Thixotropic Gel → Panels, Lipids, TSH, CMP, PSA
• Green / Heparin → Electrolytes, Coag Studies, Platelet Count
• Lavender / EDTA → Whole Blood, CBC, WBC, Sed Rate, Hgb $A_{1c}$
• Grey / Fluoride or Oxalate → Glucose, GGT, ETOH
• Royal Blue / Heparin or EDTA → Toxicology, Dilantin, Digoxin
• Yellow / ACD → Paternity, ABO Blood Type

Study Tips

• Create color-coded flashcards pairing tube color with additive & top three tests.
• Practice writing the order of draw from memory daily.
• Relate additive mechanism to test rationale (e.g., fluoride stops glycolysis → preserves glucose).
• Rehearse real-world scenarios: “STAT potassium? → Green tube.” ; “Prenatal HIV panel? → SST.”