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Practical work #7. Calculation of leukocytes
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Practical work #7. Calculation of leukocytes

Leukocytes or white bodies have many variations and perform different protective high-specified functions. That’s why the amount of leukocytes varies during a short period of time in the healthy man (from 4•109/l to 9•109/l).

Dilution of the blood for the calculation of leukocytes is better to be performed with the help of melanger.

 

In clinics other methods of blood dilutions are also used. For example, 0,4 ml of 3% solution of acetic acid, which is painted with methylene-blue is put in the dry tube, then 0,02 ml of blood is added. Blood gets by any pipette with graduation. It is important to keep correlation 1:20.

Figure 6.6- Melanger for leukocytes

Acetic acid is needed in calculation of leukocytes to cause hae molysis of erythrocytes, and methylene-blue – for the contrast of leukocytes nuclei, so for the convenience of calculation.

Objectives: to calculate the amount of leukocytes in Goryaev’s camera.

Requirements for work: microscope, lighter, Goryaev’s camera, covering glass, melanger for leukocytes, holder with tube, 3% solution of acetic acid, 96% ethanol, cotton wool.

To prepare Goryaev’s camera for work:

perform defatting with alcohol and dry the camera and the covering glass;

lap the covering glass to the camera;

find the net under the large enlargement.

Prepare blood for work:

fill the capillary till the mark of 0,5; clean the opening from blood with the cotton wool;

don’t release the blood from the capillary and fill it with 3% solution of acetic acid till the mark of 11;

mix the solution with blood in the ampule of mixing tube (there is a tiny red ball to ease the mixing process). Blood will be diluted in 20 times.

To fill camera with blood:

blow first two drops of solution out of the capillary on the cotton wool;

the next drop from the ampule solution is put in the camera. Put the tip of the melager on the edge of camera near the covering glass and blow it out accurately. Solution will go under the covering glass into the camera and will fill it. Wait for 1-2 minutes for leukocytes to sedimentate on the bottom of the camera.

Count the amount of the leukocytes:

count the amount of erythrocytes in 100 large quadrates. For better accuracy calculation should be performed on the whole area of net, starting from the left edge.

calculate the amount of erythrocytes in 1 mkl of blood by formula:

L = (a • 4000 • 20) / 1 • 100 • 16

Where L – the amount of leukocytes in 1 mkl;

a – the amount of leukocytes in 100 large quadrates of net;

100 – the amount of large quadrates;

16 – the amount of small quadrates;

20 – the degree of blood dilution;

1/4000 mm3 – the volume of 1 small quadrate

There is a simplified formula:

L = (a:2) • 102

- to find the number of leukocytes in 1 l of blood

L • 106

Recommendations for writing down the results:

Draw down the melanger for leukocytes.

Count the amount of leukocytes in 100 quadrates of net.

Calculate the number of leukocytes in 1 ml and in 1 l of blood.

Answer following questions in conclusion:

Is the amount of leukocytes normal in examined blood and what does it testify?



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