physiology lab 2, manual rbcs count

manual RBCs count

Introduction

Erythrocyte (RBCs) are biconcave in appearance Have no nucleus

The interior of the cell contain red pigment called Hemoglobin which is responsible for red color of blood.

Life span 120 days

Introduction

The function of erythrocytes to combine with O2 & to lesser extent CO2 & transport them through blood vessels.

Introduction

Normal range in healthy adult male has about 4.5-6 million RBCs/mm³

healthy adult female has about 4-5.5 million RBCs/mm³

Principle of the test

In this experiment you are going to determine the # of RBCs in mm³, and because this # is very large, dilute a sample with an Isotonic Solution (Hayem’s solution), then multiply the counted # by correction factor.

Material

1. Blood sample (EDTA anticoagulated blood or capillary blood)

2. RBCs diluting pipette3. Diluting solution (Hayemes solution)4. Hemocytometer5. microscope

RBCs diluting pipette

The Hemacytometer

]0.25mm

]0.20mm

Methodology

With a safety bulb draw blood up to 0.5 marks on RBCs pipette and complete to 101 with RBCs diluting solution.

Mix for 2-3 minute.

Charge hemacytometer

Discard the first 4-5 drops

Place tip of the pipette at the edge central platform

Then let the hemacytometer to stand on the bench for 3-5 minute.

Count and calculate:

Calculation

Number of RBCs/mm³=counted cells in 5 squares(80 small squares)

x diluting factor x volume correction factor.

Calculation

The dilution factor= total volume/sample volume = 101-1/0.5 =200 Volume correction factor= Desired volume/counted volume in 5 squares 1 mm³/ counted volume

Calculation

counted volume =The total volume of the 5 Medium

squares== Volume x number squares= (width x length x depth )x 5= (1/5mm x 1/5mm x 1/10 mm) x 5

=1/4000mm³

Calculation

Total # of small squares= 5 x 16=80 Then The total volume of the 5

Medium squares=1/4000 x 80=1/50 Volume correction factor=

1/(1/50)=50

Calculation

Example: If total # of RBCs in 5 medium

squares is 423 Then the # of RBCs in 1mm³= 423 x

200 x 50 = 4,230,000

Calculation

Number of RBCs/mm³= counted cells in 5 m square x diluting factor x volume correction factor.

Number of cells/mm³= n x200 x 50 = n x 1000

Significance of the test

The purpose of a RBCs count is to determine the # of ciculating RBCs per mm³ of blood

RBCs carry O2 to all tissues; thus a drastic reduction in the RBCs count will cause immediate reduction in the available O2.

Significance of the test

Anemia: a decrease in number of RBCs below normal range

Polycythemia: Increase in number of RBCs above normal range

Physiological Polycythemia: (up to 8 million cells/mm³) due to:

Age: at birth RBCs count is 8-10 million cells/mm³ High altitude

Significance of the test

Pathological Polycythemia: due to Primary Polycythemia: RBCs over 14

million cells/ mm³, occur in bone marrow malignancy.

Secondary Polycythemia: RBCs over 8 million cells/ mm³,due to:

Respiratory disease Heart disease Chronic carbon dioxide poisoning

Sources of error

1. Flooding of chamber with excess sample

2. Failing to count all the cells in the squares or conversely including artifacts in the count.