Order of Draw in Phlebotomy: Tube Colors, Additives and Why It Matters
By the OQMPS team ·
The order of draw is the sequence phlebotomists fill blood collection tubes in when they draw several from one venipuncture. Each tube holds a different additive, and the order keeps an additive from one tube from getting into the next and skewing the results. It is a core part of phlebotomy training, and you use it on every multi-tube draw.
The order of draw, as set by the Clinical and Laboratory Standards Institute (CLSI) for venipuncture:
- Blood culture bottles
- Light blue: sodium citrate
- Red or gold: serum tubes, with or without clot activator or gel
- Green: heparin, with or without gel
- Lavender or pink: EDTA
- Gray: sodium fluoride (glucose)
Always follow your own facility’s protocol where it differs.
In this guide
Order of draw chart
Most U.S. labs use the colors below, but manufacturers differ a little, so check the label: it is the additive, not the color, that decides where a tube goes.
| # | Tube | Additive and what it does | Example tests | Inversions |
|---|---|---|---|---|
| 1 | Blood culture bottles (or yellow SPS tube) | Culture medium. Grows any bacteria in the blood, so it must stay sterile. | Blood cultures | 8 to 10 (SPS tube) |
| 2 | Light blue | Sodium citrate. Stops clotting so clotting tests can be run on the plasma. | PT/INR, aPTT, D-dimer, fibrinogen | 3 to 4 |
| 3 | Red, gold or red-and-gray (SST) | None or clot activator; SSTs add a gel separator. Lets the blood clot so the serum can be separated. | Metabolic panels, lipid profile, liver tests, hepatitis and HIV screening | 5 (plastic tubes) |
| 4 | Green or light green (PST) | Sodium or lithium heparin. Stops clotting for tests run on plasma, often when results are needed fast. | STAT chemistry and troponin (light green); ammonia, ionized calcium, lithium (green) | 8 to 10 |
| 5 | Lavender, purple or pink | EDTA. Stops clotting and preserves the blood cells for counting. | CBC, hemoglobin A1C, ESR (lavender); blood type and antibody screen (pink) | 8 to 10 |
| 6 | Gray | Sodium fluoride, often with potassium oxalate. Stops the cells using up glucose while the sample waits. | Fasting and timed glucose, lactate, blood alcohol | 8 to 10 |
Example tests are from one hospital laboratory’s 2025 guide; which tube a test goes in varies between labs, so your lab’s test catalog has the final word. Specialty tubes, such as royal blue for trace elements, are drawn in the position that matches their additive.
How to remember the order of draw
A popular mnemonic is “Stop Light Red, Stay Put, Green Light Go”:
- Stop: sterile, the blood cultures
- Light: light blue
- Red: red
- Stay: SST, the gold serum separator
- Put: PST, the light green plasma separator
- Green: green
- Light: lavender
- Go: gray
Red and SST are both serum tubes, so some labs list the SST first; either way, they come after light blue and before green.
It also helps to remember the logic rather than the colors. The sterile sample goes first; the clotting test comes before anything that could affect clotting; serum tubes come before the anticoagulant tubes; and the tubes whose additives cause the most trouble elsewhere, EDTA and fluoride, come last.
Why the order of draw matters
When the needle passes from one tube to the next, a trace of the last tube’s contents can travel with it. Each additive does a specific job, and in the wrong tube it interferes with the test.
EDTA is the best-known example. One hospital laboratory’s procedure warns that EDTA “can yield false magnesium and calcium results” and “can falsely elevate potassium results” if it gets into a serum or heparin sample. That is because the potassium salt of EDTA adds potassium, while EDTA binds calcium and magnesium.
Blood cultures go first for a different reason: they detect bacteria, so they are drawn before anything else “to reduce contamination,” as VCU Health’s guide puts it.
How much does the order of draw really matter?
Less than its reputation, under ideal conditions, but still enough to follow. In a 2021 study of 388 outpatients drawn in the “wrong” order with a closed collection system, researchers found no EDTA contamination at all. They did still see potassium results shift between tubes, and concluded that order of draw “has significant influence on the potassium results” regardless.
A 2017 case report in the same journal found real EDTA contamination in practice, and noted that while some evidence suggests the order no longer matters under ideal conditions, the European Federation of Clinical Chemistry and Laboratory Medicine’s working group on the pre-analytical phase still recommends continuing to advise it. Real draws are not always ideal, which is why the rule stays.
Butterflies, syringes and fingersticks
Winged sets (butterflies) and the discard tube
The tubing on a butterfly holds air. If a light blue tube is the first one you need, that air would underfill it. So BD and CLSI-based lab procedures say to draw a discard tube first: a plain (no additive) or another citrate tube, just enough to fill the tubing, and not sent for testing. It does not need to be full.
Fill the light blue tube to the line
Clotting tests depend on the ratio of blood to citrate. As one hospital’s procedure puts it, “Partially filled blue top tubes will be rejected.”
Syringe draws
The same order applies when you draw with a syringe: fill the tubes in the same sequence, using a blood transfer device.
Fingersticks and heelsticks: a different order
Skin punctures follow their own order (CLSI GP42). Blood clots quickly as it pools on the skin, so the anticoagulant tubes come first:
- EDTA (lavender), first, so the cell counts are accurate
- Other additive tubes, such as heparin
- Serum tubes last
Some labs collect capillary blood gases and blood smears before these. As always, your lab’s procedure decides.
Mixing the tubes
Every tube with an additive needs mixing straight after it is filled. Invert it gently, turning it upside down and back, the number of times in the chart above. Do not shake it. Under-mixing an anticoagulant tube can leave small clots that make the sample unusable.
Common order of draw mistakes
- Going by color alone. Read the label: the additive decides the position, and specialty tubes slot in by additive.
- Skipping the discard tube when a butterfly is used and the light blue tube comes first, which leaves it underfilled.
- Underfilling the light blue tube. Clotting tests need the right blood-to-citrate ratio, and labs reject short tubes.
- Using the venipuncture order for a fingerstick. Skin punctures take EDTA first.
- Shaking instead of inverting, or not mixing at all.
Studying for your certification exam? The order of draw is part of the phlebotomy curriculum at OQMPS. See what to expect on the phlebotomy certification exam.
Learn it hands-on
OQMPS phlebotomy classes are 3 days in class plus self-paced home study, in Arizona, Oregon, Texas, Utah and Washington. Can’t get to a campus? Take the course online.
Frequently asked questions
What is the correct order of draw in phlebotomy?
For venipuncture, the CLSI order is: blood culture bottles, light blue (sodium citrate), red or gold serum tubes, green (heparin), lavender or pink (EDTA), and gray (sodium fluoride). Follow your facility's protocol where it differs.
Why are blood cultures drawn first?
Blood cultures detect bacteria, so they must stay sterile. Drawing them before any other tube reduces the chance of contamination.
Which tube is drawn last?
Of the standard tubes, gray (sodium fluoride, for glucose) is drawn last. Some labs draw certain specialty tubes after it; follow your lab's list.
Is the order of draw different for a fingerstick?
Yes. For skin punctures, CLSI's capillary standard puts EDTA (lavender) first, then other additive tubes, with serum tubes last, because blood collected from the skin starts clotting quickly.
What is a discard tube?
A tube drawn first, and not tested, when you use a butterfly and a light blue citrate tube is the first one needed. It fills the air in the tubing so the citrate tube gets the right amount of blood. It can be a plain tube or another citrate tube and does not need to be full.
Does the order of draw apply to syringe draws?
Yes. Fill the tubes from the syringe in the same order, using a transfer device.
Sources
- Laboratory procedure, Order of Draw (Southeast Iowa Regional Medical Center), citing CLSI GP41 7th ed. (2017) and GP42-A6
- BD Vacutainer Order of Draw for Multiple Tube Collections (hosted by Texas DSHS)
- VCU Health Department of Pathology, Order of Draw (rev. 2/2025)
- UCHealth, Order of Draw procedure SC003.1
- University of Iowa Health Care, Phlebotomy Tubes and Order of Draw
- Ercan et al., Order of draw of blood samples affect potassium results without K-EDTA contamination during routine workflow, Biochemia Medica (2021)
- Cornes, Case report of unexplained hypocalcaemia in a slightly haemolysed sample, Biochemia Medica (2017)