How to determine bleeding | clotting | prothrombin| APTT| time

                               

          How to determine bleeding and clotting times

 This article discusses different methods for determining bleeding and clotting times. Learn about the tests, procedures, indications, and more.

Table of contents

1.	Definition
2	Apparatus and Reagents
3.	Procedures Clotting factors
4.	Normal values
5.	Alteration in normal values

 

Keywords: Duke's method | "Ivy" method | Von Willebrand's disease | Purpura | Bernard Soulier disease | Disseminated intravascular coagulation (DIC) | Wright's method | Lee and White method | Capillary glass tube method | Haemophilia| 

It is essential to determine the bleeding and clotting time in bleeding disorders before surgical operations and many other conditions.

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                        Determination of the bleeding time

Principle: If the finger is deeply pricked, the time taken between the appearance of blood and the cessation of bleeding is bleeding time.

Apparatus and Reagents:- 

  1. Apparatus for sterile finger prick, i.e., pricking needle, cotton swab, spirit.

  2. Filter paper.

 3. Stopwatch.

Procedure: Duke 1990 measured the time for bleeding to stop from a standardized wound in the ear lobe; therefore, this method is Duke's. Many new techniques are present even now, and it is the widely used method.

Duke's method: The tip of the left ring finger is cleaned with a spirit swab and dried. To get free-flowing blood, a bold prick 3mm deep is made with a sterilized special pricking needle or lancet. A stopwatch is started, and time is noted with the help of the stopwatch. Blood is mopped up every 15 seconds without applying pressure on the finger until the bleeding stops. The number of blood spots on the filter paper determines the bleeding time.  

Calculation of the bleeding time:

We got seven blood spots on the filter paper when blood was mopped up every 15 seconds.

Then bleeding time is =7x15 =105 seconds.

105 seconds=1minute45 seconds.

Result: 

1 minute 45 seconds,

Inference: It is within the standard limit.

Another method to determine the bleeding time is 

The "Ivy" method (Haemostasis bleeding time) is more reliable than Duke's method. It requires a sphygmomanometer.

 Discussion: A normal bleeding time reflects sufficient platelet number, functions, and normal microvasculature.

The average value is -1 to 3 minutes.

It is prolonged in    1. In the case of purpura, only B.T. is prolonged, and C.T. is normal.

                                  2. In Von Willebrand's disease, both B.T. and C.T. are prolonged.

                        3. Disseminated intravascular coagulation (DIC)

                        4. Bernard Soulier disease.

Precaution: Antiseptic and aseptic measures must be followed.

                   

    Give a bold prick so that blood flows freely.

                                                Determination of clotting time 

Principle:- If the finger is deeply pricked, the time between the appearance of blood and clot formation is clotting time.

Apparatus and Reagent : 

  1. Apparatus for sterile finger prick, i.e., pricking needle, cotton swab, spirit

  2. Fine capillary glass tube of about 15cm. 

  3. Stopwatch.

Procedure :

 Methods: Two methods are commonly used:

1. The capillary glass tube method is Wright's, and 2. Lee and White method.

The capillary glass tube method is frequently used.

First, the fingertip of the left ring is cleaned with a spirit swab and dried. Next, a bold prick is made with a sterilized pricking needle to get free-flowing blood. A stopwatch is started, and time is noted. Next, blood is drawn into a capillary glass tube by putting one of its open ends into the blood. By capillary action, blood enters the capillary tube and fills it. The capillary glass tube is then kept between the palms of both hands for one minute to keep it at body temperature.

After one minute, the tube is removed. A small portion of the capillary tube is broken at intervals of 15 seconds until a thread of clotted blood of about 5mm in length appears between the two pieces of the capillary glass tube. The time between the appearance of the blood on the skin and the thread of the blood clot gives the clotting time.

Calculation of the time :

After eight pieces of the capillary glass tube, the thread appears. The capillary glass tube was broken at a regular interval of 15 seconds.

Result :

Then clotting time is =8x15 =120 seconds+1minute=3 minutes. 

(120 seconds=2minutes.)

Inference: It is within the standard limit.

Discussion:  Normal value is -2 to 4 minute

In Haemophilia, only the C.T. is prolonged, Liver disease, Jaundice, esp. obstructive jaundice.

 In Von Willebrand's disease, both B.T. and C.T. are prolonged.

Shortened in:  Physiological condition: After a meal, menstruation, and parturition.

Pathological condition: After hemorrhage, after blood transfusion, and febrile states.

 Precautions: Antiseptic and aseptic measures must be used.

                   

     Give a bold prick so that blood flows freely.

                       Time must be noted accurately.        

                        The capillary glass tube should be about 15 cm long.

Indication of BT and CT

1. Family history of bleeding disorders.

2. History suggestive of hemophilia or other bleeding disease- spontaneous bleeding, frequent and prolonged bleeding even after minor injuries.

3. Before any minor or major surgery.

4. Before performing a biopsy from internal organs like the liver and kidney.

5. Before starting anticoagulant drugs, assess the effects of the drugs.

Factors influencing BT

1. Condition of blood vessels.

2. Nature of wound.

3. Number and functional conditions of thrombocytes.

4. Ambient temperature -BT is short during cold climates.

Factors influencing CT

1. Cotting factors presence or absence.

2. Ambient temperature prolongs the clotting time.

3. It depends on the contact surface.

Tests are present to estimate the quantities of clotting factors. The prothrombin test, which estimates the amount of prothrombin, is called prothrombin time. We will delve into prothrombin time.

Prothrombin: The average plasma prothrombin is 30-40 mg/dl.

Bleeding may occur when the prothrombin level decreases below 20% of normal.

Prolonged PT regular PT is 15-20 seconds.

1. Deficiency of prothrombin factor II,

2. It suggests a deficiency of factors V, VII, and X.

3. Liver diseases.

4. Vitamin K deficiency.

Multiple tests are present to estimate the quantities of clotting factors. 

The prothrombin test—prothrombin time—is commonly used to determine prothrombin quantity and functions. We will delve into prothrombin time.

Prothrombin time examines extrinsic pathways and common pathways of the coagulation mechanism.

Introduction

Prothrombin time is the time it takes for the plasma to form a clot in the presence of thromboplastin and calcium ions.

You take 0.2 ml of 3.8% sodium citrate solution in a 2 ml. syringe.

A tourniquet is applied over the upper arm to make the antecubital vein prominent. The skin over the vein is cleaned with a cotton swab soaked in rectified spirit and allowed to dry. With strict aseptic and antiseptic precautions, the needle punctures the vein, and up to 2 ml of blood is drawn in the syringe.

After drawing the blood, the tourniquet is removed quickly, and the puncture wound is pressed with a cotton swab soaked in rectified spirit for a few minutes to stop the bleeding.

Transfer the blood mixed with the sodium citrate solution into a centrifuge tube. Then, centrifuge in an electrical centrifuge machine for 30 minutes at 3000 rpm.

After that, 0.2 ml citrated plasma is taken from the centrifuged tube into a small test tube, and 0.2 ml of thromboplastin suspension is added. The tube is placed in a water bath to maintain a temperature of 37 degrees Celcius. Now, 0.2 ml warm up to 37 degrees Celcius, and calcium chloride solution is added to the tube.

As soon as calcium chloride solution is added, time is noted till a clot appears in the tube. This is Prothrombin time.

Explanation: Calcium ions, along with other coagulation factors and tissue factors, are essential for blood coagulation. A sodium citrate solution removes the calcium ions, and citrated plasma is free from calcium ions.

In the second step, calcium chloride solution and thromboplastin suspension are added to provide calcium ions and thromboplastin, respectively.

Prothrombin: The average plasma prothrombin is 30-40 mg/dl.

Bleeding may occur when the prothrombin level decreases below 20% of normal.

Prolonged PT

The average prothrombin time is 10-25 seconds.

A decreased prothrombin time is rare, but if present, it may increase the chances of thromboembolism.

Prolonged prothrombin time:

1. Deficiency of prothrombin factor II,

2. It suggests a deficiency of factors V, VII, and X.

3. Liver diseases.

4. Vitamin K deficiency—due to any cause, such as reduced supply or decreased absorption from the gastrointestinal tract.

Partial thromboplastin test (PTT) =activated partial thromboplastin test(aPTT)=Thromboplastin test.

Its historical name is Kaolin-Cephalin clotting time (KCCT) because Kaolin and Cephalin were used in this test.

It was initially used as a lab reagent to determine blood clotting quality, but it was later recognized as a clotting factor.

This test reveals the coagulation mechanism's internal and common pathways.

Thromboplatin is factor III. It is present mainly in the brain, lungs, liver, platelets, and other organs.

This factor is present in the cell membrane and has two components: Phospholipids and tissue factors.

It depends on the subject's age, sex, health condition, and laboratory equipment.

Typical range is 25-35 seconds. In infants, the upper limit is 50 seconds.

Short APPT suggests an increased risk of thromboembolism.

Increased APPT indicates some bleeding conditions and the use of certain drugs.

The indications are as of Prothrombin time or determination of clotting time. In addition, it is performed when a problem with one or more clotting factors is suspected. This test diagnoses von Willibrand's disease.

APPT indirectly measures functions of factors I, II, VIII, X, XI, and XII.

Functions of factors VII AND XIII are not detected in this test.

This is known as the partial thromboplastin test due to the absence of tissue factor in the reaction mixture.

In a blood sample at 37 degrees Celsius, a coagulation activator, e.g., silica, kaolin or ellagic acid) is added, and the time is recorded when the clot is formed. This is APTT.

The thrombin generation test (TGT) is the thrombin generation assay (TGA), which is the Global coagulation assay (GCA).

Take home message 

 If the finger is deeply pricked, the time taken between the appearance of blood and the cessation of bleeding is bleeding time. The average value is 1-3 minutes and is determined by Duke's method.

If the finger is deeply pricked, the time between the appearance of blood and clot formation is clotting time. In the lab, it was determined that the capillary glass tube method was Wright's. The average value is 2-4 minutes.

 FAQ :

Q. What is bleeding time? 

A. If the finger is deeply pricked, the time between the appearance of blood and the cessation of bleeding is bleeding time.

Q. Give an example where bleeding time is prolonged?

A. Bleeding time is prolonged in purpura.

Q. What is clotting time?

A. If the finger is deeply pricked, the time between the appearance of blood and clot formation is clotting time.

Q. Give an example where clotting time is prolonged?

A. Clotting time is prolonged in the case of hemophilia.

Q. Give an example where both bleeding and clotting times are prolonged?

A. In von Willebrand's disease, both B.T. and C.T. are prolonged.

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 Hashtags: Duke's method # "Ivy" method (Haemostasis bleeding time)# Willebrand'snd's disease # Purpura# Bernard Soulier disease# Disseminated intravascular coagulation (DICWright'sht's method # Lee and White method# Capillary glass tube method # Haemophilia#

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