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Vacutainer- Needle, Holder and Types of Blood Collection Tubes

The vacutainer system is the set of vacutainer needle, holder, and blood collection tube. As the name implies, a vacutainer is a complete system of drawing blood from the patient to the vacuum blood collection tube. These tubes consist of different kinds of preservatives or the anti-coagulant for carrying out different kinds of blood tests. Based on the constituents present within the tubes, the tube can mainly be EDTA tube, serum tube, heparin tube, sodium fluoride tube, coagulation tube, ESR tube, etc.

We will be discussing the workings, uses, and color-coding of these kinds of tubes in this blog post.

Vacutainer Needle

Vacutainer Needle Parts
Vacutainer Needle Parts

A vacutainer needle is a type of needle, that is used to draw blood from the patient.

As shown in the figure, the major components of the vacutainer needles are the bevel, shaft, threaded tub, and rubber sleeve covering the needle.

The sizes of the needle generally range from 1 to 1.5 inches with the gauge value ranging from 19 to 22.

A single needle can be used to draw blood from a patient multiple times. However, the same needle should not be used to draw a sample from different patients. Else, there can be chances of cross-contamination.

Vacutainer Holder

A vacutainer holder is a holder that holds the vacutainer tubes. It helps in the process of blood collection. It also protects from the exposure of communicable diseases to some extend.

The holder is made up of disposable plastic. It is reusable in nature. Besides that, you can even sterilize it with the help of an autoclave.

How to use Vacutainer holder

To know the proper use of the vacutainer holder, go through the video shown below.

Vacutainer Blood Collection Tube

Vacutainer tube is a type of test tube that has got a colored rubber cap on top of it. This rubber cap helps to create a vacuum within the tube. It is either made up of plastic or glass.

The vacutainer tube is sterile within and generally consists of certain constituents. The constituents help to preserve the specimen for a longer period of time. Thus, this helps to carry out different types of blood tests.

Vacutainer Color Code

The color coding can be on the basis of an International Standard or on European Standard. The color-coding does not depend upon the cap of the vacutainer tube. Rather it depends upon the constituents present within the tubes.

Vacutainer Tube Color Code (International Standard) Color Code (European Standard)
EDTA  Tube Lavender Cap Red Cap
Serum Tube (Without gel separator)

Serum Tube (With gel separator)

Red Cap

Yellow Cap

White Cap

Brown Cap

ESR Tube Black Cap Purple Cap
Sodium Citrate Tube (Coagulation Tube) Blue Cap Green Cap
Heparin Tube Green Cap Orange Cap
Sodium Fluoride Tube Gray Cap Yellow Cap

Vacutainer Types

(Which vacutainer tube for what tests? )

On the basis of constituents present within the tube, or on the basis of the function of the tube, the tube can be classified into the following types.

1. EDTA Tube (Ethylene Diamine Tetra Acetic Acid)

According to the international standard, the EDTA tube has got a lavender cap. Similarly, on the basis of European standards, it has got a red cap.

Working of the EDTA Tube

EDTA acts as an anticoagulant in a vacutainer blood collection tube. Thus, it prevents clot formation. The EDTA blood sample can easily last for at least 4 hours at room temperature and for few days inside the freezer.  Also, it helps to preserve the morphology of the blood cells.

Coagulation is possible due to the binding nature of the EDTA to the calcium present in the blood.  Calcium is responsible for the coagulation of the blood. However, it has a higher binding affinity to the EDTA. Hence, the calcium-EDTA complex prevents the calcium molecule to bind with the blood factors. Thus, the blood will not clot. This process is also irreversible in nature.

Instead of normal EDTA, K2EDTA or K3EDTA may also be present in the tube.

EDTA Tube Use

(EDTA Tube Function)

The EDTA tube can be appliable for a number of applications. Some of the applications of the EDTA tubes are as follows.

However, the EDTA blood is not useful for the measurement of the divalent ions such as calcium and magnesium. Also, it is not useful for enzymatic tests.

2. Serum Tube (Serum Separating Tube)

According to the international standard, the serum tube without the gel separator has got a red cap. Similarly, on the basis of European standards, it has got a white cap. However, if the tube has got the serum gel separator, the serum tube has a yellow cap according to the international standard. Similarly, on the basis of the European standards, it has got the brown cap.

Serum Tube Composition

The serum tube consists of silica. This silica is responsible for the clotting of the blood. Thus, it helps to separate the serum from the other factors.

Silica consists of negative charges onto the surface. This helps in the activation of factor twelve, also known as the Hageman factor.  The Hageman factor helps in the activation of the intrinsic pathway for coagulation.

Silica is a major constituent of glass tubes. But nowadays, plastic blood collection tubes are mostly in use. In such a case, the inner layer of the plastic tube is coated with silica or contains silica-coated beads.

As stated above, the serum tube can also consist of the serum gel separator. The specific gravity of the serum gel lies between that of the serum and the blood components. So, during the process of centrifugation, the serum gel separates the serum from the blood components. This helps to prevent the serum from cross-contamination from the blood components.

Serum Separator Tube Gel Composition

The most commonly used gel composition is given below.

  • 55 to 83% (w / w) of n-butyl acrylate (NBA)
  • 9 to 37% (w / w) of 2-ethylhexyl acrylate (2-HA)
  • 2.0 to 3.5% (w / w) of silicon dioxide
  • 2 to 10% (w / w) of styrene monomer (SM)
  • 0.3 to 1.0% (w / w) of Rheology modifiers

Serum Separating Tube Uses

Some of the applications of the serum separating tubes are as follows.

  • Urea and Electrolytes (Na, K, Iron, Cl, and Ca)
  • C- Reactive Protein (CRP)
  • Liver Function Tests such as  SGPT, SGOT,  Bilirubin, Albumin, Total Protein, and ALP
  • Bone Profile Tests such as Calcium, Phosphate, Albumin, and ALP
  • Tumour Markers such as AFP, PSA, CEA, CA-125, and CA19-9
  • Toxicology Tests such as Ethanol, Cannabis, and Opiates
  • Drug Levels Tests such as PCM, Salicylate, and Aspirin
  • Immunology Tests such as Immunology Ig, and Rheumatoid Factor.

3. Coagulation Tube/ Sodium Citrate Tube

According to international standards, the coagulation tube has got a blue cap. Similarly, on the basis of the European standard, it has got the green cap.

Since the coagulation tube consists of sodium citrate, it is also called the citrate tube. Just like the EDTA, the sodium citrate binds with the calcium to form the calcium-citrate molecule. But, it has got a lower affinity in comparison to that of the EDTA. Two calcium ions can remove three citrate ions. So by adding the required amount of calcium, the coagulated blood can be reversed.

Sodium Citrate Tube Uses

It is used in the coagulation screen.

  • B.T:- Bleeding Time
  • P.T:- Prothrombin Time
  • aPTT:- Activated Partial Thromboplastin Time
  • D-dimer test:- For thrombosis
  • INR:- International Normalized Ratio

Sodium Citrate Tube Ratio

The sodium citrate tube ratio is defined as the ratio of sodium citrate to the blood sample. The sodium citrate tube must be exactly 1: 9. That means, 0.1 ml of sodium citrate should be mixed with 0. 9 ml of the blood sample.  In conclusion, the coagulation tube should neither be overfilled nor be underfilled.

If the concentration of the citrate is higher in the blood sample, then it will remove more calcium from the blood. Thus, the coagulation time will increase. Similarly, if the concentration is lower in the blood sample, a significant amount of calcium will be present in the blood. Hence, the blood will clot quickly.

Sodium Citrate Tube Vs EDTA

Both sodium citrate and the EDTA tube works by removing calcium from the blood sample. The coagulation within the EDTA tube is irreversible. But, in the case of sodium citrate tubes, the coagulation process is reversible. Hence by adding calcium to the tube, the coagulation can be triggered in a controlled environment. In other words, it allows for an analysis of the coagulation factors in the blood.

4. Heparin Tube

The heparin tube is mainly of two types. One is the sodium heparin tube while the other one is the lithium heparin tube.

According to international standards, both of the heparin tubes have got a green cap. Similarly, on the basis of European standards, they have got the orange cap.

Heparin tube consists of heparin in it. Heparin acts as an anticoagulant substance that binds with the inactivated antithrombin leading to its activation. The activated antithrombin binds with the factor Xa causing the reduction of the formation of thrombin from prothrombin. Hence, this prevents the formation of fibrin from fibrinogen. Fibrin is the cause of blood coagulation.

Heparin Tube Tests

Some of the tests performed with the help of the heparin tube are stated below.

  • Ammonia
  • Insulin
  • Renin and Aldosterone

(Note:- Sodium heparin tube is not suitable for the measurement of electrolytes. Because the sodium present in the heparin tube, can interfere with the test and give an elevated value of sodium in the blood sample. In such a case lithium heparin tube is useful.

A lithium heparin tube is not suitable for the measurement of lithium levels in the blood. In such a case, a sodium heparin tube is useful. )

EDTA Vs Heparin

The function of the heparin tube is similar to that of the EDTA tube. They are both responsible for the inhibition of the coagulation process. So, the EDTA and the heparin tube are interchangeable in most cases. EDTA works by removing calcium from the blood. On the other hand, heparin operates by activating the antithrombin. This, in turn, inhibits the formation of thrombin.

The advantage of the EDTA tube over the heparin tube is that it can also inhibit the calcium-dependent enzymes. Thus, the stability of the samples becomes high. So, the parameters can be analyzed for a longer period of time.

However, the EDTA affects cell functions. Thus in the case of the functional analysis, the EDTA tube is not useful.  In such a case, a heparin tube is preferred.

5. Sodium Fluoride Tube

According to international standards, a sodium fluoride tube has got a gray cap. Similarly, it has got the yellow cap, according to the European standard.

It is used to measure glucose or lactate levels in the blood.

Glycolysis can affect the measurement of blood glucose levels. Glycolysis causes a decrease in the blood glucose level and an increase in the lactate level. So, to inhibit the glycolysis process, fluorine ions are added. Other inhibitors such as EDTA, calcium oxalate, or potassium oxalate may also be used in a trace amount.

6. ESR Tube

ESR tube can be easily identified due to its thin diameter. According to the international standard, it has got a black cap. Similarly, on the basis of European standards, it has got a purple cap.

ESR, also known as the Erythrocyte Sedimentation Rate, gives useful guidelines for any kind of inflammatory process. During the measurement, the collected blood is stored at room temperature for a certain period of time. The cells will begin to sediment at the bottom of the tube.

The ESR tube is a thin tube that has leveling in it. In other words, it is similar to the measuring cylinder, where the readings are in the mm scale. As time goes by, the RBC begins to settle at the base. With the help of the measuring scale, the rate of sedimentation is measured. The displacement of the RBC is measured for a period of an hour

Since the ESR is only possible in anticoagulated blood, anticoagulated citrate is also added in the ESR tube.

ESR Range

ESR Range
ESR Range

There is no exact value for the ESR. The value can alter from one person to another. Factors that mainly affect the ESR value are age and sex. The value of ESR in female is generally higher than that of the male. Similarly, the value increases with an increase in age. Besides that, geographical location may also affect the value.

As stated before, the table shown here is just an approximate value.  Besides this, the value of ESR for newborns ranges from 0 to 2 mm/hour. Also, the value of ESR for neonatal to puberty ranges from 3 to 13 mm/hour.

ESR Function

Any deviation of the ESR value from the normal range can be an indication of inflammation. However, the result that we obtain from this method may not be much reliable. Several other factors can alter the value. For example, an increase in the ESR value can be due to pregnancy, anemia, autoimmune disorders, etc. Similarly, a decrease in the ESR value can be due to sickle cell anemia, low plasma protein, leukemia, etc.

In conclusion, the ESR value may be relevant only when the difference with the reference value becomes too high.

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