Session 3: Blood Counts and Films

Question 1

Give examples of why a full blood count might be deviant other than disease. What else can go wrong?

Answer 1

Mix up of specimen collection
Wrong bottle
Pooling samples
Poor technique
Wrong delivery method
Specimen delayed or not even delivered
Wrong test requested or performed
Incorrect clinical details
Variability

Question 2

What should you do if you get an unexpected or abnormal result of the full blood count?

Answer 2

Repeat the test.

Question 3

How is a full blood count performed?

Answer 3

It’s done using a pink top EDTA sample.
This requires k-EDTA which is an anticoagulant that chelates calcium ions.
This stop the blood from clotting.
If it would clot the test will be rendered useless.

Question 4

How is haemoglobin measured? (And most of the blood cells)

Answer 4

By spectrophotometry. Determined by lysis of red cells followed by conversion of haemoglobin to a stable form.

It is measured in g/L (concentration)

Question 5

Explain spectrophotometry.

Answer 5

The amount of light absorbed by sample is proportional to amount of absorbent compound within it.

Question 6

What does spectrophotometry depend on?

Answer 6

The wavelength of the light shone through.

Question 7

Explain flow cytometry.

Answer 7

A hydrodynamic focussing where single file line of cells are put through a light beam.
What you measure is the impedance and how the cell scatters.
The more scatter from a cell the bigger it is.

Question 8

There are different types of scattering. What does forward scatter measure? What does side scatter measure?

Answer 8

Forward: size
Side scatter: Mono/polymorphnuclear and intracellular complexity.

Scatter can also measure myeloperoxidase activity.

Question 9

How is white blood cell count measured?

Answer 9

Automated cell counting AFTER the red blood cells have been lysed in the sample.

Question 10

How is red blood cell count measured?

Answer 10

Automated cell counting. This is before the lysis.

Question 11

Since red blood cell count is measured before the red cells are lysed, wouldn’t the white blood cell count show up as well?

Answer 11

Yes they would and they do. However the amount of white blood cells is so small compared to red blood cells so it doesn’t significantly affect red blood cell count.

Question 12

What is haematocrit?

Answer 12

The fraction of whole blood volume that consists of red blood cells this can be called packed cell volume (PCV) as well.

Question 13

What is haematocrit used for?

Answer 13

Anaemia but more often polycythemia.

Question 14

What is mean cell volume (MCV)?

Answer 14

The average volume of red cells measured in femtolitres (10^-15).

Question 15

What is mean cell volume used for?

Answer 15

To determine whether and anaemia is microcytic or macrocytic.

Question 16

What is mean cell haemoglobin?

Answer 16

The average amount of haemoglobin protein measured in femtolitres in an individual red blood cell.

Question 17

How is mean cell haemoglobin calculated?

Answer 17

Dividing haemoglobin concentration in a given volume by the number of red cells in that same volume.
Hb/MCV
Haemoglobin/Mean cell volume

Question 18

How is platelet count measured?

Answer 18

By the same principle as red and white cells. Since platelets are much smaller than red and white blood cells they can easily be distinguished.

Question 19

How is reticulocyte count measured?

Answer 19

Reticulocytes are counted using special stains or fluorescent dyes which bind to ribosomal RNA.

Question 20

Why is rRNA stained for reticulocyte count?

Answer 20

Because immature red blood cells (reticulocytes) still have rRNA. Mature red blood cells do not have any rRNA.

Question 21

Why is reticulocyte count measured?

Answer 21

Usually to evaluate different kinds of anaemia.

Question 22

How is a blood film (peripheral blood smear) performed?

Answer 22

By placing a drop of blood at one end of a slide which is the thinly dispersed using a spread slide to obtain a monolayer of cells which can be easily viewed under the microscope.
The sample is the air dried and fixed in methanol before staining in order to distinguish different cell types.

Question 23

What is a blood film used for?

Answer 23

Assessment of causes for a low or high RBC/WBC count.
Looking for blood parasites such as malaria or to check for abnormalities such as sickle cell anaemia, spherocytosis and thrombotic thrombocytopenia purpura.
Also to look for the presence of Howell-Jolly bodies in order to see if the patient is hyposplenic.