Erythrocyte Count

• Is the number of erythrocytes per micro litter of blood.

• Normal ranges:–Male 5 - 5.5 106 /µL – Female 4.5- 5 106/µL– New born 6-8 106/µL

• Erythrocyte count increased in case of polycythemia and decreased in anemia.

Erythrocyte Count• Principle:• In order to facilitate RBCs count a specified volume of

blood is diluted with a specified volume of isotonic fluid..

Diluting fluid:• One of the following solutions may be used:

1. Isotonic saline: – 0.85% sodium chloride (NaCl) in distilled

water.2. Hayam’s solution:

– Sodium Sulphate 5 g.– Sodium Chloride 1 g.– Mercuric Chloride 0.5g.– Distilled Water 200ml

• Sample:– Whole blood using EDTA or heparin as

anticoagulant. Capillary blood may also be used.• Equipments:

1. (Pipettes) used one of the following: • (RBCs) pipette• Micropipette –20l is the desired volume.

2. Improved Neubauer chamber with the cover slips.

3. Conventional light microscope.

4. Clean gauze.

DILUTION FACTORS For RBC counting

Blood is filled till mark 0.5 and Hayem’s fluid is then filled till mark 101.Both are thoroughly mixed and then few drops are discarded which contain just the diluting fluid in the stem. Thus,

1 portion out of 101 is discarded. So ,0.5 part of blood is in 100 parts of fluid or,

1 part of blood is mixed in 200 parts of fluidThus, dilution factor for RBC counting is 200.

RBC COUNTING

• Total no. of RBCs in 5 secondary squares = X• Mean No. of RBCs in 5 secondary squares = X/ 5• X/ 5 x 25 = No. of RBCs in the primary sq (1 mm2) of

diluted blood• X/ 5 x 25 x200 = No. of RBCs in the primary sq (1 mm2)

of pateint blood• Because the depth between the cover and slide was only

0.1 mm• X/ 5 x 25 x200 x 10 = No. of RBCs in the primary sq (1

mm3) (= 1 µl of blood)

= X x 10,000/mm³ or µL

X is the number of RBCs in 5 secondary squares

Coagulation timeDefinition :-It is the time needed for formation of a hard clot or it is the time elapses between appearance of bleeding and formation of clot .Material :-- Clean dry glass slide .,- stop watch ,- water bath 37º c- syringe,- alcohol , cotton .Procedure :-*obtain blood in the syringe and start stop watch since appearance of the blood in the syringe .* Put a drop of the blood on the slide*the two slide are placed on a warm plate at 37ºc Every 30 seconds put a needle in the middle of the blood drop and try to elevate it *blood co aggulation start when fibrin threads. Appear attached to the needlesNormal : 4-10 minutes .

Clotting Time - Slide Method (Intrinsic)•The surface of the glass tube initiates the clotting process. This test is sensitive to the factors involved in the intrinsic pathway•The expected range for clotting time is 4-10 min.

Haemostasis or blood clotting

Extrinsic way Intrinsic way

It is a blood test that looks at how fast small blood vessels in the skin close to stop bleedingPurpose•To assess overall hemostatic function (platelet response to injury and functional capacity of vasoconstriction). • To detect congenital and acquired platelet function disorders. Patient preparation •Normal = 3-5 min

Bleeding time

• It is the time needed for bleeding from small wound to stop without coagulation .

• Material :-• - sterile lancets .- stop watch . - sphygmomanometer .• - glass slide .- Ethyl alcohol .•  Procedure :-

• Method • *A sphygmomanometer cuff is placed on the upper arm and

inflated to 40mm Hg .• *A sterilized area of forearm without visible superficial veins is

selected and is pierced by sterilized lancet in 3 points .• *wipe the blood every 30 sec from the 3 points alternatively until

no blood comes to filter paper .•Normal : up to 5minutes