Angeles University Foundation

College of Allied Medical Professions

WHITE BLOOD CELL COUNT

Written Report
Midterms

In partial fulfillment of the requirements in

Hematology Laboratory

Submitted to:
Mrs. Analyn Navarro, RMT
Mrs. Eloisa Singian, RMT

Submitted by:
Guevarra, Sonny
Mungcal, Lilibeth
Sarmiento, Norilie Mae
Tayag, Joseph

Group 8

BSMT 3a

January 28, 2009

INTRODUCTION
 White blood cells (WBCs), or leukocytes, are cells of the immune system
defending the body against both infectious disease and foreign materials. Five
different and diverse types of leukocytes exist, namely: the neutrophil, eosinophil,
basophil, lymphocyte, and the monocyte, but they are all produced and derived
from a multipotent cell in the bone marrow known as a hematopoietic stem cell.
Leukocytes are found throughout the body, including the blood and lymphatic
system. The name "white blood cell" is derived from the fact that after
centrifugation of a blood sample, the white cells are found in the buffy coat, a thin,
typically white layer of nucleated cells between the sedimented red blood cells and
the blood plasma. The scientific term leukocyte directly reflects this description,
derived from Greek leukos (white), and kytos (cell).
The number of WBCs in the blood is often an indicator of disease. There are
normally between 4×109 and 1.1×1010/L white blood cells in a litre of blood, making
up approximately 1% of blood in a healthy adult. An increase in the number of
leukocytes over the upper limits is called leukocytosis, and a decrease below the
lower limit is called leukopenia.
Quantitative evaluation of the formed elements in the blood is an important
factor in making a diagnosis. This is termed as hemocytometry. For white blood
cells, it is specifically known as the white blood cell count. A WBC count is normally
ordered as part of the complete blood count (CBC). In some circumstances, a WBC
count may be ordered to monitor recovery from illness. There are different methods
in the numerical estimation of the blood cells. They are: the turbimetric method,
microscopic method and the automated method.
The turbimetric method is based on the assumption that the more turbid a
solution, the more cells are present in the blood. This method is obsolete and very
erroneous. The automated method makes use of calibrated machines that uses that
makes use of light beams or voltage pulses in order to accurately count the cells.
The method used for counting white blood cells in the experiment is the
microscopic method. Cells are counted under the microscope using the following
materials:
a. Counting chamber
b. Pipettes (specifically the WBC pipette)
c. Diluting fluid

a. The Counting Chamber

The most commonly used type of counting chamber is the improved
Neubauer. The counting chamber (hemocytometer) has 2 ruled areas on each
surface, each containing a square divided into large 9 squares, each with an area of
1mm square. The depth of the counting chamber is 0.1mm.
 Figure 1: The Counting Chamber showing the areas for counting the WBC (marked
as W)
and the WBC’s that are counted and not counted (colored in
green).

The four corner large squares (W) are used in counting the White Blood Cells.
Each square has an area of 1 sq. mm, and are subdivided into 16 smaller squares.
The inverted rule of L (marked in red) is used in counting the cells. All cells that are
inside the square and those that fall on the lines of the rule of L are counted.

b. The WBC Diluting Pipette

The Thoma cell counting pipette is a calibrated glass pipet with a bulb for a
diluting
chamber. There are two types of pipets, one for WBC counting (characterized by a
clear or white mixing button in the mixing chamber) and the second for RBC
counting
(identified by the red mixing button in the diluting chamber). Each has a pipet stem

with calibration marks. Both pipets do not measure in mLs, but in parts. Each pipet
is designed to give a specific dilution.

The Thoma WBC pipet can dilute from 1:10 to 1:100. Most WBC pipets
contain ten
calibration marks designated as 0.1 to 1.0. A final calibration mark is located on the
opposite side of the bulb (designated by 11). The volume in the stem is 10 times
less
than that of the bulb. Blood, pipetted to the 0.5 mark, then diluted to the 11 mark
provides a 1:20 dilution. Note the dilutions possible using the WBC pipet and
pipetting blood (or any body fluid) to the 0.1 mark and diluted to the 11
mark give a 1:100 dilution.

THE WBC THOMA DILUTING

PIPETTE
[Type a quote from the
document or the summary of
an interesting point. You can
position the text box
anywhere in the document.
Use the Text Box Tools tab to
change the formatting of the
pull quote text box.]

The

Trenner pipet differs from the Thoma pipet in the way the stem is joined to the
mixing bulb. The stem inserts into the bulb so that the end is flat, polished, and
at right zangles to the longitudinal axis. The means that blood can be drawn into
the stem by capillary action and will fill the stem, automatically stopping at the end
of the stem. Each Trenner pipet is calibrated to dilute to a designated volume.

THE TRENNER DILUTING

PIPETTE
 A body fluid is drawn at the designated mark on the stem and diluted to the
11 mark on the opposite side of the bulb gives the following dilutions.

Mark on the stem WBC pipette dilution

0.1
1:100
0.2
1:50
0.3
1:33
0.4
1:25
0.5
1:20
0.6
1:17
0.7
1:14
0.8
1:12
0.9
1:11
1.0 1:10

The stem contains pure diluting fluid and that is the reason why it is always
discarded as the first few drops and the reason why we subtract 1 from 11.
Therefore 11-1= 10 in which it is constant in the WBC bulb.

c.The Diluting FLuids

In WBC counts, the diluting fluids aim to lessen the number of blood cells and
lyse cells that are not needed in the count (red blood cells are lysed but not the
nucleated RBC).

1. 2-3% Glacial Acetic Acid

Composition:
Glacial acetic acid 2 or 3ml
Distilled Water 97 or 98 ml
Gentian Violet (1% aqueous) 1ml
 2. 1% Hydrochloric Acid
Composition:
1N Hydrochloric Acid 1ml
Distilled water 99ml

3. Tuerk’s solution
Composition:
Glacial acetic acid 2ml
Distilled water 97ml
Methyl violet 1 drop

d. Procedure

Suck the blood Suck diluting Shake Discard the

to the 0.5 mark fluid to the 11 pipette to first few
of the pipette. mark. mix. drops

Compute for the Count the WBC in Charge the

WCC the 4 corner large counting chamber
squares

e. Calculations

The formula for the White Cell Count is:

WCC = WBC counted x DCF x VCF

Where:

WCC = White cell count

DCF = Dilution Common Factor

= _Volume in the Bulb (10 constant for WBC)__

Volume of Blood Used (variable)

VCF = Volume Correction Factor (2.5)

= _______1 __________
 0.4 cu.mm (constant)

*0.4 cu. mm is obtained by multiplying the volume of

1 corner square which is 0.1 cu. mm to 4 (total corner
squares
• Short-cut method if the dilution factor is 20

WBC in thousand/cu. mm = WBC counted x 50

f. Things to remember
1. Over charging will lead to the to the decrease in wbc count because cells will
fall into the moat of the counting chamber.
2. It is important to allow the counting chamber to stand for a 3 minutes to after
charging to allow cells to settle down.
3. Always discard 2-4 drops of the mixture before charging
4. To check if the cells are uniformly distributed, subtract the highest cells
counted from the lowest cell counted (cells counted in each of the 4 corner
large square), the difference should not exceed 15. If it exceeded 15, this
means the cells are not uniformly distributed and needs to be recharged.
5. The corrected WBC count should be done when there is a high WBC count
and more than 10 nucleated RBC per 100 RBC in the blood smear.

CWCC = _ uncorrected WCC x 100___

# of nucleated RBC + 100

g. Significance of the results

The normal values for the WBC count in 6,000-11,000/ cu.mm. An elevated
number of white blood cells is called leukocytosis. This can result from bacterial
infections, inflammation, leukemia, trauma, intense exercise, or stress.

It is important to say that high white blood cell count would be considered normal in
certain situations:
• Pregnancy in the final month and labor may be associated with increased WBC
levels.
• Spleen removal could grant persistent mild to moderate increased WBC count.
• Normal newborns and infants have higher WBC counts than adults
A decreased WBC count is called leukopenia. It can result from many
different situations, such as chemotherapy, radiation therapy, or diseases of the
immune system.
EXPERIMENTAL RESULTS

Name of Patient: Norilie Mae Sarmiento

Age: 19 yrs. old

Date/ Time of Extraction: January 7, 2010

Type of Specimen: Diluted blood

WBC count (Traditional unit): 2,600 /cu. mm 3,300/

cu.mm

WBC count (SI unit) : 2.6x109/ L 3.3x109 /

L

Final Result (average of 2 counts): 2.95x109/L (SI)

2,950cu.mm (Trad)

Normal Values: 6,000-11,000/cu. mm

Interpretation: Low wbc count

Calculations:

Counting Chamber 1
Counting Chamber 2

W1 = 13
W2 = 12
W3 = 16 W1 = 18
W4 = 11 W2 = 13
W3 = 17
Total = 52 wbc’s W4 = 18

WCC = WBC counted x DCF x VCF Total = 66 wbc’s

= 52 x 20 x 2.5
= 2600/cu.mm WCC = WBC counted x DCF x VCF
= 66 x 20 x 2.5
 = 3,300/cu.mm

QUESTIONS FOR RESEARCH

1. Enumerate 5 sources of error or conditions and identify their

corresponding effect on WBC count:

Sources of Error Effect on WCC

Overflow of the fluid in the gutter Decreased

Failure to discard fluid before charging Decreased

Trapped bubbles mistaken for WBC Increased

Clotted blood Decreased

Unlysed nucleated RBC Increased

2. How are abnormal white blood cells treated?

 The treatment for leukocytosis, or a high white blood cell count, depends
upon the cause. Only an investigation by your doctor can identify the
appropriate treatment. For example, if the cause of the elevated WCC in
bacterial, then it is treated by antibiotics and bone marrow transplant for
leukemia.

Treatment of leukopenia is also based upon the underlying cause, severity,

and the presence of associated infections or symptoms as well as the overall
health status of the patient. Obviously, treatment must also be directed toward
any underlying disease process. Treatments that directly address leukopenia
may include (note that all of these treatments may not be appropriate in a given
setting):
• administration of white blood cells growth factors (such as recombinant
granulocyte colony-stimulating factor (G-CSF, filgrastim) in some cases of
severe neutropenia;
• granulocyte transfusions; or
• corticosteroid therapy or intravenous immune globulin for some cases of
immune-mediated neutropenia.

1. Identify diseases associated with the abnormal results of WBC

count:

Increased WBC count Decreased WBC count

Leukemia Systemic Lupus Erythematosus

Inflammatory (ex. Rheumatoid Bone Marrow failure

arthritis)

UPDATES
References

http://www.labtestsonline.org/understanding/analytes/wbc/test.html

http://www.nlm.nih.gov/medlineplus/ency/article/003643.htm

http://www.labtestsonline.org/understanding/analytes/wbc/test.html#when

http://www.clt.astate.edu/wwilliam/hem_i_intro.htm

http://www.healthandage.com/high-white-blood-cell-count-why-is-my-white-blood-
cell-count-too-high#Treatments_for_High_White_Blood_Cell_Count