1) What is bleeding time?

Whenever an injury or cut occurs in skin, bleeding occurs for a while but it usually stops
without any external help within few minutes.

"The time elapse between the skin puncture and arrest of bleeding is called bleeding time"

It is also a crude test of hemostasis (the arrest of bleeding). It indicated the function of
platelets in blood.

2) What is the normal value of bleeding time?

2-6 minutes

3) What is the role of platelets to stop bleeding?

Platelets adhere to the endothelial surface of traumatized blood vessel and release
various chemical substances that cause vasoconstriction of the blood vessel and also
form platelet plug, which is then converted to a clot through complement cascade
system (intrinsic and extrinsic pathway).

Platelets have glycoprotein coat that help in adhesion of these platelets to the injured
vessel.

Have growth factors that promote growth and multiplication of damaged vascular
endothelial cells.

They have enzyme system to synthesize prostaglandins

They have fibrin stabilizing factor that causes polymerization of fibrin monomers to form
fibrin threads.

Its contains large amounts of phospholipids that activate multiple stages of blood
clotting process.

4) What are the clinical conditions in which bleeding is prolonged?

o Deficiency of platelets (thrombocytopenia).

o Deficiency of vitamin K and calcium.
o Decrease prothrombin, factor 7, 9 and 10 caused by vitamin K deficiency.
o Hemophilia due to deficiency of factor 8.
o Inadequate formation of fibrinogen.
o Disease of liver such as hepatitis, cirrhosis and acute yellow atrophy can develop
sever tendency to bleed as almost all the clotting factors are formed by liver.
o Von Willebrand disease.
o Bernard-Soulier syndrome.
o Treatment with anti-coagulant drugs such heparin and warfarin.
5) Define and difference between Duke's and Ivy's method?
Duke's method
o The Duke method involves puncture
with a lancet or special needle
preferably on the earlobe or fingertip.
o The prick is about 3-4 mm deep.
o The test ceases when bleeding
ceases.
o The patient then wipes the blood
every 30 seconds with a filter paper.
o The usual time is about 1-3 minutes.
Ivy's method
o Utilizing an incision on the ventral
side of the forearm.
o A standard-sized incision is made
around 10 mm long and 1 mm deep.
o The time from when the incision is
made until all bleeding has stopped
is measured and is called the
bleeding time.
o Every 30 seconds, filter paper or a
paper towel is used to draw off the
blood.
o A normal value is less than 9 and half
minutes.
1) What is clotting time?
"The time between collection of blood, after puncture and formation of thread."
Or
The time elapses between the withdrawal of blood and clot formation."

3) Why blood does not clot within the blood vessels normally?
o In the blood stream, the anti coagulants normally predominate to the pro-
coagulants so that blood does not coagulate while it is circulating in the
blood vessels.
o The smoothness of the endothelial cell surface, which prevents contact
activation of the intrinsic clotting system.
o A layer of glycol-calyx on the endothelium, which repels platelets and
clotting factors, thereby preventing activation of clotting
o Thrombomodulin- thrombin complex, which slows the clotting and also
activates a plasma protein C that acts as anti-coagulant by inactivating
factors V and VIII
o The anti-coagulants naturally present in body are calcium-free
thrombokinase and heparin and warfarin, protein C.

4) What initiates the process of clotting?

An injury or trauma to the blood vessel, (due to which the smoothness and glyco-
calyx layer of the endothelium is lost) which activates both factor XII and platelets,
initiates the process of clotting.
The pro-coagulants over-ride the anti-coagulants.

5) What are the naturally occurring anti-coagulants in blood?

o Heparin
o Protein C
o Fibrin fibers
o Anti-thrombin III
o Anti-thrombin-Heparin co-factor

6) Which anti-coagulants are generally used in laboratory?

o Heparin
o Various substance that decrease the concentration of calcium ions in the
blood can also be used as anti-coagulants for example a soluble oxalate
compound, which causes precipitation of calcium oxalate from plasma,
thereby decreasing the ionic calcium level so much that blood coagulation is
blocked.
o Any substance that deionizes the blood calcium such as sodium. Ammonium
or potassium citrate.
o Coumarins such as warfarin as anti-coagulants.
 o The blood-collecting test tubes are siliconized, which delays clotting for
about an hour.

7) What is the role of calcium ions in the mechanism of coagulation?

Except for the first two steps in the intrinsic pathway, calcium ions are required for
promotion and acceleration of all the blood-clotting reactions.

8) Name the clinical conditions in which the clotting time Is prolonged

o Include a decreased plasma fibrinogen level, thrombocytopenia,
dysfibrinogenemia, paraproteinemia, heparin contamination, elevated levels
of fibrin degradation products, and liver failure.
o Muscles and joint bleeding indicates coagulation defect.
o Prolong bleeding from superficial cuts, GIT, hemorrhage or menorrhagia
indicates a failure of platelets or possibly the presence of vluf disease.
o Recurrent bleeds at a single site suggest a local structural abnormality.
o Operational surgery as dental surgery, tooth extract, tonsillectomy,
circumcision accounts bleeding but continuous bleeding even after surgery,
indicates deficiency of platelets.

9) What is the significance of determining clotting time?

Clinical significance

Pathologic changes

A) Increased values
• Christmas disease,
• congenital afibrinogemia,
• congenital dysfibrinogenemia,
• factor II deficiency,
• factor V deficiency,
• factor XI deficiency,
• factor XII deficiency,
• heat stroke,
• hemophilia,
• leishmaniasis,
• SLE,
• smallpox,
• toxic effects of venom,
• von Willebrand’s disease.

B) Decreased values
• eclampsia.

10) What are the general principles to stop bleeding of a shed blood?
11) Is there any other method than capillary tube for determination of CT?
What is anemia?

Deficiency of hemoglobin is known as anemia

1) What are the causes of anemia?

Anemia can be due to following reasons

Decrease erythrocytes in blood, which may be due to malfunctioning in erythropoisis or

inhibition or un-sufficient release of erythropoietin or any damage to the bone marrow.

Iron deficiency due to decrease iron intake in diet or mal absorption

Excessive blood loss from the body.

Increased hemolysis of RBCs such as in hemolytic anemia or systemic lupus arythmatosis.

Pregnancy state.

Alcoholism

Kidney disorders as its releases erythropoietin.

HB

1) What is hemoglobin?
Hb is the iron-containing oxygen-transport metalloprotein in the red blood cells.

2) What is the function of hemoglobin?

Hb binds O2 molecules with it and transports it from lungs to the cells and carries
CO2 from cells back to the lungs.

3) What are the normal types of hemoglobin?

HbA ………………….. 2 alpha + 2 beta (in adults)

HbA2 ………………… 2 alpha + 2 delta

HbF …………………. 2 alpha + 2 gamma (present in fetus)

4) What is the normal concentration of Hb in males and females?

Male………………………. 14-16 gm/dl

Female………………….. 12-14 gm/dl

5) What is anemia?

Deficiency of hemoglobin is termed as anemia.

6) Name the types of anemia.
1) Microcytic hypochromic anemia.
 Iron deficiency anemia
2) Normocytic/ Normochromic
 Hemolytic anemia
 Hereditary spherocytosis
 Sickle cell anemia
 Aplastic anemia
3) Macrocytic anemia.
 Megaloblastic anemia.
 Pernicious anemia.

7) How much O2 is carried out by 1gm of hemoglobin?

1gm of hemoglobin carries 1.34 ml of O2

8) What is the effect of hydrochloric acid on hemoglobin?

Hb is converted into acid hematin by action of HCl.

9) What is the name of brown pigment which is formed?

Acid hematin.

1) Define ESR?
ESR: Abbreviation for erythrocyte sedimentation rate, a blood test that detects and
monitors inflammation in the body. It measures the rate at which red blood cells (RBCs)
in a test tube separate from blood serum over time, becoming sediment in the bottom
of the test tube.

2) What is the principle of this experiment?

Well mixed whole blood containing 3.8% trisodium citrate as an anticoagulant is
placed in a calibrated Westergen tube and is allowed to stand vertically and
undisturbed for exactly one hour. The distance in mm through which red cells
settle during 1 hour period is reported as ESR.

3) What are the clinical conditions in which ESR increases?

ESR is greatly enhanced by those conditions which increase the density of RBCs,
thus increasing rouleaux formation.
These are:-
 Hyperfibrinogenemia (increased fibrinogen level)

 Immune hypergammaglobulinemia (increased antibodies during immune response)

 Hypoalbuminemia (low plasma albumin)

 Anemia
 Inflammatory bowel disease

 Acute pancreatitis

 Cholecystitis

 Peritonitis

 Hepatitis

 Kawasaki disease

 Wagener's granulomtosis

 Bechet's syndrome

 Churg Strauss syndrome

 Hypersensitivity vasculitis

 Urticarial vasculitis

 Systemic lupus erythematosus

 Rheumatoid arthritis

 Pyelonephritis

 Hemolytic Uremic Syndrome

 Acute glomerulonephritis

 Chronic glomerulonephritis with renal failure

 Nephrosis

 pregnancy

4) What are the clinical conditions in which ESR decreases?

Sickle cell disease

Anisocytosis
Spherocytosis
Acanthocytosis
Polycythemia
Extreme leukocytosis
Microcytosis
Hypofibrinogenemia
Hypogammaglobulinemia
Dysproteinemia
 Hyperviscosity state
Glucocorticosteroids
Cachexia
Congestive heart failure

5) What are the factors on which ESR depends?

 Erythrocytes:- rouleaux formation
 Composition of plasma
 Technical factors: - it is important that ESR tube in held absolutely vertical,
tilting of tube greatly increases ESR.

6) Why the normal value of ESR is different in male and female?

The ESR range is greater in females as compared to males because it is
dependent on rouleaux formation.

Less num of RBCs increase inter-erythrocytic collision increase rouleaux

formation increase ESR

Greater number of RBCs decrease inter-erythrocytic collision decrease rouleaux

formation decrease ESR

1) What is the normal WBC count?

4000-11000/ ul

2) How will you identify WBC pipette?

WBC pipette is identified by white bead within the bulb

3) What is the significance of using Turk's fluid?

It is a blood diluting fluid that causes hemolysis of RBCs and stains the nucleus of
leukocyte so that WBCs can be well observed under microscope.

4) What is the function of bead in the pipette?

It helps in mixing of the blood with the Turk's fluid within the bulb portion.

5) What do you mean by 1:20 times dilution

It means 20 parts of the Turk's fluid diluting 1 part of the blood sample.

6) How much is the depth of Neubaur Chamber?

7) What major precautions you will follow in charging the chamber?
The charging chamber and cover slip should be free of dust.
Before charging, blow out the fluid present at the end of the pipette.
No air bubble should be present in the counting chamber.
 8) In what clinical conditions the TLC decreases
Pancytopenia, typhoid fever, malaria.

9) In what clinical conditions TLC increases

Infections, leukemia's.

10) What are the physiological variations for TLC?

1. after exercise

2. during emotional outbursts

3. after meals

4. Pregnancy- Return to normal after delivery

5. at birth

6. Diurnal variation

What are the physiological variations in the number of RBCs?

1. Diurnal -

2. Age -

3. Sex -

4. High altitude -

5. Muscular exercise -

- Polycythemia (Erythrocytosis)

- Anemia (Erythropenia)

Less during night, minimum in early morning, gradually increases during day

In newborn, high count is seen

More in males

What are the conditions leading to polycythemia?

Polycythemia is a physiological increase in RBCs, especially in people who live at high altitudes, as
there is less oxygen in the environment at higher altitudes.

How much is the life span of RBCs

120 days

What is the normal count of RBCs?

Male 4.5-5.5 millions/ mm3

Female 3.9-4.5 million/mm3

Why RBCs are biconcave in shape?

The biconcave shape of the RBC helps them in circulation especially when passing through thin
capillaries.

What are the functions of RBCs?

They help in oxygen transport from lungs to the cells.

Help in acid-base buffering.

Contains large quantity of an enzyme Carbonic anhydrase which catalyze reversible reactions
between CO2 and H2O

How are RBCs formed?

They are formed in the bone marrow form the pleuripotent hematopoitic stem cell (PHSC). The
erythropoisis is stimulated by a hormone "erythropoietin"

What are the causes of anemia?

Anemia can be due to following reasons

Decrease erythrocytes in blood, which may be due to malfunctioning in erythropoisis or

inhibition or un-sufficient release of erythropoietin or any damage to the bone marrow.

Iron deficiency due to decrease iron intake in diet or mal absorption

Excessive blood loss from the body.

Increased hemolysis of RBCs such as in hemolytic anemia or systemic lupus arythmatosis.

Pregnancy state.

Alcoholism

Kidney disorders as its releases erythropoietin.