Reviewer ( Hema Lab )
Topic 6: RBC COUNT
Red Blood Cell
Compromises almost 45% of the human blood
It is responsible for the transport of oxygen and nutrients throughout the body
Principle
To dilute the blood in an isotonic fluid without damaging or lysing the
cells Computation
# of cells counted x Correction for dilution x correction of volume x 10^6
Where in:
Dilution: 1 : 200
Correction for volume: 50
Normal Values
Male
4.2 - 5.4 x 10^12 / L
Female
3.6 - 5.0 x 10^12 / L
Newborn
5.5 - 6.5 x 10^12 / L
Topic 7:
Hemoglobinometry
Hemoglobin
Is an iron containing protein molecule found in Red Blood Cells which transport
oxygen from the lungs to the body’s tissue
In Adult Human Body there are 3 kinds of normal hemoglobin molecules
HbA1 ( A2 B2 )
96 - 98%
HbA2 ( A2 D2 )
1.5 - 3.2%
Hb F ( A2 Y2 )
0.5 - 0.8%
Can be tested manually or using n automated machine
It is important to diagnose the patient for possible anemia or
polycythemia Hemoglobincyanide Method
Is the internationally recommended for determining the
hemoglobin concentration of the blood
Reference method for Hb estimation cause:
All Hb forms except sulphaemoglobin are estimated
Highly reliable and stable reagents are available
The reagent is sensitive to light
It contains cyanide, making it highly toxin
The method can be easily standardized
It takes an hour to convert in heavy smokers
However, the value increased is not clinically significant
Principle
Blood is diluted in Drabkin’s Solution…
Test Sample
Venous or capillary blood collected into an EDTA tube
Sources of Error
Turbid samples due to increased wbc or platelet may cause false increase result
This can be corrected by centrifugation of the samples and testing the
supernatant
Another reason for turbidity is lipemic specimen
To correct this, add 0.1 of Patient’s plasma to 5 ml of reagent and use the
solution as blank
HbS and HbC can be resistant to hemolysis and cause turbidity
To correct this, add 1:2 solution of diluted sample water, then multiply
the
standard cure by 2
Addition of potassium carbonate are used to correct abnormal globulins
Abnormal Hemoglobin in Adults
Carboxylhemoglobin
Hemoglobin + carbon monoxide
Methemoglobin
Hemoglobin in which ferrous iron are oxidize to ferric states
Sulfhemoglobin
Irreversible formation of from sulphonamides and aromatic
amines
Normal Ranges
Male
13 - 17 g / dl
Female
11.5 - 16.5 g / dl
Newborns and infant
14 - 22 g / dl
Topic 8: Hematocrit
Determination
Hematocrit
The packed red cells of red blood cells
Constitute to the volume of red blood cells over the volume of whole
blood Principle
Constitute of the use of centrifuge o separate red blood cells from whole blood
to its maximum packaging to measure the percentage expressed red blood cells
over the whole blood
Normal Values
Neonates
45 - 60%
1 year old above
24 - 44%
Women
36 - 48%
Men
40 - 55%
Methods
Micro hematocrit
Macro hematocrit
Rarely used for it is time and sample consuming
Make used of wintrobe tube nd centrifuged at the
2000 - 2300 from 30 minutes
Topic 9: Peripheral Blood Smear
Blood Smear
Can be a key to different picture in the blood film of the patient
Types of Blood Smear
Cover Glass
smear
More even distribution of WBC but they are harder to master, too small for
automated strainers, and easily broken
Wedge Smear
Most commonly used blood smear in the
laboratory Components
The feathered Edge ( Head )
This is the end of the blood
smear Monolayer ( Body )
This is the optimal area for examination of cells
Base of the smear ( Tall )
These areas are usually too thick to evaluate cells
under high power
Spun Smear
More easily handled and labelled with properly distributed WBC and RBC
Buffy Coat Smear
Buffy coat is used instead of whole blood in smear preparation
This is done to verify if there is leukopenia or thrombocytopenia
Thick Blood Smear
This is usually done in identification of blood parasites such as malaria,
babesia..
Stains used in hematology
laboratory Romanovsky Stains
Contains methylene blue and eosin
Wright’s Stain
Methylene Blue and eosin azure
Giemsa Stain
Eosin Y and azure
B Wright’s - Giemsa Stain
Leishman Stain
Jenner Stain
May - Grunwald Stain
Methanol
Fixative or stabilizer
Counting Method
Cross Sectional / crenellation
Longitudinal Method
Battlement Method
Things to Examine in a Blood Smear
Check first the condition of blood smear if this is well -
made Perform WBC differential count
Scan for abnormal blood cells
Count 100 WBC in the smear, categorizing each based on its identification
If there are presence of nRBC, do not include them in the 100 WBC’s that you
counted, instead, take note of how many nRBC’s you encountered while
completing he 100 WBC, then report it as numbers of nRBC’s / 100 WBC
Examine the RBC morphology
Presence of poikilocytes
Presence of inclusions and abnormalities
Presence of immature RBC’s
Take note the size and pallor of
RBC’s Rouleaux and Agglutination
Polychromatophilia
Examine the morphology and number of platelets
Computation for absolute differential count
Absolute # cells / L = % of cell type in differential x WBC / L
Topic 10: Erythrocyte Sedimentation Rate
Erthrocyte Sedimentation Rate
Is a non specific test that indicates changes In plasma protein concentrations
due to infection or injury
It is the rate at which the red blood cells settles in a tube over a given amount
of time
Factors affecting
ESR Plasma
Protein
Changes in plasma protein occur rapidly following tissue injury or in
response of inflammation
RBC Size and Number
The size and number of RBC’s that show alterations in their biconcavity,
usually do not increase the ESR rate
Technical Factors
Tilted ESR tube
ESR tube should always be in perpendicular position
Temperature
Temperature higher than room temperature accelerates
sedimentation
Vibration
Can reduce the rate of erythrocyte sedimentation
Methods of Measuring ESR
Westergren method
Recommended
Principles
It measures the sedimentation of red cells after 1 hour in
an open ended tube mounted vertically on a stand
Wintrobe and Landsberg Method
