The order of draw is the fixed sequence for filling blood tubes, starting with blood culture bottles, then sodium citrate, serum, heparin, EDTA and fluoride. The sequence stops additives from one tube reaching the next. Follow your trust chart, because UK charts differ.
Additives travel through backflow, direct transfer or pouring between tubes. EDTA in a serum sample raises potassium and lowers calcium. Clot activator in a citrate tube shortens clotting times. The lab then reports a number the patient never had. Closed vacuum systems show little carryover.
Draw blood with a method you trust. A wrong sequence leads to a repeat draw, and the patient feels the second needle. Royal Open College’s Phlebotomy and Venipuncture Training course covers vein selection, vacutainer handling, contamination prevention and sample labelling. The course is CPD learning, not a regulated qualification, so your employer signs off practical skills.
Follow your site’s chart, which usually matches the CLSI sequence of cultures, citrate, serum, heparin, EDTA and fluoride. Carryover is a real risk, though studies find the effect most clearly with syringes and open collection. UK charts differ in colours and in where citrate sits.
The order of draw is the sequence for filling tubes during one venepuncture. CLSI lists one order for syringe, tube holder and pre-evacuated systems. The 2025 edition, PRE02, replaced GP41. Its published change list names no change to the tube sequence.
The CLSI order of draw
Step | Tube | Example colour | Why this tube sits here |
1 | Blood culture bottles | Varies | Helps protect sterile samples from contamination by tube stoppers or additives |
2 | Sodium citrate | Light blue | Helps prevent clot activator carryover into coagulation tests |
3 | Serum, with or without clot activator or gel | Red or gold | Comes before EDTA, which can falsely raise potassium |
4 | Heparin | Green | Comes before EDTA, which can interfere with electrolyte results |
5 | EDTA | Lavender or pink | Placed late because potassium EDTA can raise potassium and lower calcium results |
6 | Sodium fluoride and potassium oxalate | Grey | Usually placed last to reduce additive carryover into other tubes |
Closure colours are examples only. Makers use different colour systems, so read the additive name on every tube. Some UK charts add tubes outside the six steps, such as cross-match EDTA, ESR citrate and trace element tubes. Match the additive, never the colour.
Only blood cultures, plain glass serum tubes and plastic serum tubes without clot activator go ahead of citrate. Clot activator carried into a citrate tube shortens PT, so clotting tests need this protection.
A discard tube follows its own rule, covered in the butterfly section below.
The order limits additive carryover, the transfer of anticoagulant or clot activator from one tube into the next. Carryover changes results from the second tube. Laboratory staff then work from numbers misstating the patient’s condition, and patients face repeat draws or unneeded treatment.
Studies propose three routes, and none is proven for every collection setup. Closed vacuum systems keep tubes capped and avoid syringe contact. Open methods expose blood to staff and surfaces. Knowing the routes shows where a draw is weakest.
In UK investigations, EDTA contamination of serum samples traced almost always to syringe tip or needle tip contact, or to pouring blood between tubes. Staff holding tubes upside down during filling is a suspected route, though no study has tested the idea.
EDTA contamination raises potassium and lowers calcium, magnesium, zinc and iron, depending on the test method. PT and aPTT lengthen. Clot activator moves clotting times the other way and shortens PT. A falsely high potassium worries laboratories most, since patients have received emergency treatment they did not need.
In a 2017 UK audit, the median laboratory logged 19 samples with EDTA, 8.5 from a drip arm and 1 with citrate over three months. Inpatient wards supplied 80% of the contaminated samples. Only 52 replied, many in part, so treat the figures as a guide.
Guidance still recommends one order, but studies disagree on how much carryover happens with vacuum tubes. Of 11 studies in a 2021 review, seven found a negligible effect and four confirmed the need. The split follows the collection system, not the tube order.
European laboratory bodies keep the recommendation because the rule is easy to follow and ideal draw conditions are not guaranteed on every ward. A 2018 joint recommendation grades the order step 1B and covers closed systems only. Guidance and evidence sit side by side, and neither cancels the other.
Limits apply to the research. The review found few recent studies and no registered protocol. Many studies used healthy volunteers, small samples and a single tube brand, so results do not transfer to every setup. Treat the order as a low-cost safeguard, not a proven rule for every tube system.
Quick check: Which collection method shows carryover most clearly?
Answer: Open syringe methods.
Not always. UK trust charts differ in colours and in where citrate and serum sit. Among the charts reviewed, Greiner and BD sites list citrate first, while the two listing serum first appear to follow Sarstedt practice. Your own trust chart governs your practice.
Document | Nation | Date | Citrate and serum | Tube system |
NHS Lothian chart | Scotland | Date not shown | Serum, then citrate | Multifly tubing named |
North Bristol NHS Trust | England | 2014 sheet | Citrate, then serum gel | BD |
York and Scarborough SOP | England | Dec 2022 | Plain or serum, then coagulation | Sarstedt preferred |
Stockport tube guide | England | Jan 2024 | Citrate, then serum gel | Greiner |
Southern HSC Trust chart | Northern Ireland | Mar 2018 | Citrate, then serum | Greiner |
Trusts often adopt the maker’s sheet, so charts track the tube supplier. Some trusts issue a stand-alone guideline, and the Royal Cornwall Hospitals Trust guideline runs from April 2025 to April 2028. These examples cover England, Scotland and Northern Ireland. No Welsh chart appears in this review.
No UK law sets the order of draw in the documents reviewed. CLSI and tube makers publish guidance, and each trust turns guidance into protocol. One English trust requires trust training and an OSCE assessment before staff take blood, and students work under direct supervision.
Training shows in the numbers. A 2014 survey of 763,577 specimens from a Scottish hospital found higher haemolysis and EDTA contamination when trained phlebotomists did not collect the samples. New starters should ask about training and supervised practice before any solo draw.
Step 1: SYSTEM — Identify your tube system
↓
Confirm whether your workplace uses BD, Greiner, Sarstedt or another manufacturer.
Step 2: SET — Find your local order-of-draw chart
↓
Check the chart for that specific tube system in your clinical area, laboratory guide or local procedure.
Step 3: STANDARD — Compare it with the CLSI sequence
↓
Compare your local chart with the six-step CLSI order of draw and note any differences in tube order or instructions.
RESULT: Follow your local approved chart
↓
Use the order specified by your workplace or laboratory. If anything is unclear or conflicts with the reference standard, check with the laboratory team, assessor or trainer before proceeding.
Destination: The correct order of draw for your tube system and clinical setting.
System, Set, Standard is a working method, not an industry term. Run the three steps before your first solo draw and again after any change of tube supplier. Record the chart’s version and date so you know when to review.
CLSI lists the same order for syringe, tube holder and pre-evacuated tubes. Butterfly sets and capillary samples add rules of their own. Syringe transfer carries the clearest contamination risk, so use an approved transfer device and keep the standard order.
Do not pour blood from one tube into another. Tell your supervisor or the laboratory, and expect a repeat sample if a result looks wrong. Pouring is a known route for EDTA contamination, and laboratories spot trouble from result patterns such as high potassium with low calcium.
Follow four steps when you notice an error.
These are the errors to watch for.
Order errors happen. A Danish pilot recorded them in 21% of 126 observed draws, and pooled data from 16,118,499 requests show 1.99% specimen rejection, mostly from clotting, haemolysis and low volume. Neither figure is a UK rate, and the rejection data does not name the order as a cause.
Clear feedback works. After one UK laboratory added report comments on open collection and fill order, weekly EDTA cases fell from a median of 5.6 to 2.3 per 10,000 samples, a 56% drop. A potassium result that does not match the patient deserves a repeat before treatment starts.
Use a memory aid built on additive names, since closure colours differ between makers. The CLSI order runs Culture, Citrate, Serum, Heparin, EDTA, Fluoride. The first letters give C, C, S, H, E, F, so try the phrase Clean Care Starts Here, Every Fill.
In 2016, Hamilton Health Sciences in Canada gave staff colour-bead charms as a memory aid for the order. Collection errors fell afterwards, though the hospital did not establish whether the charms caused the drop.
A: No UK law sets the order of draw in the documents reviewed. CLSI and tube makers publish guidance, and your workplace turns the guidance into its own protocol. If you are new to blood collection, a course such as Phlebotomy and Venipuncture Training from Royal Open College helps you read your workplace protocol with confidence.
A: No. Pouring blood between tubes is a known route for EDTA contamination, which raises potassium and lowers calcium. Tell your supervisor or the laboratory and follow the local protocol for a repeat sample.
A: Tube makers use different closure colour systems, and trusts adopt the supplier’s chart. Greiner, BD and Sarstedt each follow their own scheme. Read the additive name on the label instead of relying on colour.
A: Employers set training and competency checks, and one English trust requires trust training and an OSCE assessment, with students under direct supervision. Competence comes from supervised practice, not from a certificate alone.
A: No general rate exists, because studies measure selected samples. Closed vacuum systems rarely show carryover, while open syringe methods do. Your laboratory holds local contamination figures and is able to share them.
A: No. The order limits carryover between tubes only. Drip arm contamination from infusion fluid and citrate from an ESR tube are separate causes, so follow your local rules for drawing near an infusion.
A: Blood cultures go first to protect sterile samples from contamination. Later tubes carry additives and non-sterile stoppers, and a culture must stay free of both. Citrate follows, then the remaining tubes in sequence.
A: CLSI PRE02 (2025) replaced GP41. Its change list names duplicate removal, alignment with other CLSI documents and updated references, and names no change to the six-step sequence. Check the edition date on any chart you use.
A: Practice varies by laboratory. In the 2017 UK audit, 96% of laboratories detected EDTA contamination from the analyte pattern, so expect a call or a repeat request. Learning how to label, handle and prepare samples for transport, which the Royal Open College course covers, helps you avoid other preventable problems.
A: No. Your employer decides competence and signs off supervised practice, and CPD accreditation is not an Ofqual regulated qualification. A CPD course such as Phlebotomy and Venipuncture Training from Royal Open College builds theory and records completed learning.
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