TRANSCRIBED GEN CAMATO

HEMATOLOGY LABORATORY
HEMATOLOGY 1 | LABORATORY

HEMATOCRIT ESR
Parallel to hemoglobin Erythrocyte Sedimentation Rate
Volume occupy by erythrocytes in the given volume of blood Rate of setting of RBC from the plasma after an addition of
Useful in determining erythrocyte indices and for calculation of a anticoagulant
blood volume 9 Very useful to establish anemia; & other disorder in blood Importance to measure rate of fall
9 MCH & MCV Hemoglobin, Hematocrit & RBC Associated with the net-surface charge of RBC with normal
Useful for total erythrocyte mass determination surface of charge negative
Useful in establishing anemia
IMPORTANCE OF ESR
METHODS FOR HEMATOCRIT Use as a good index for determination of a hidden but active
1) Macro method Use large volume of blood disease such as tuberculosis and carcinoma
a) Winthrobe method Measures the suspension stability of the RBC
Uses Winthrobe tube Measures the abnormal concentration of fibrinogen and protein
Flat bottom tube graduated at both side globulin
Fill the tube with blood sample using capillary pipet
Uses oxalated blood sample ESR METHODS
Alternative anticoagulant: EDTA 1. Winthrobe Method
Centrifuge the tube with the speed of 3,500rpm for Uses double oxalate; alternative: EDTA
30minutes Fill in the capillary pipet, stand for an hour

Calculation for % Hematocrit 2. Westergren Method * For RESEARCH purpose

Volume % of Hematocrit = Hematocrit of Packed RBC Most sensitive method of ESR determination use for serial
X 100
Hematocrit of White Blood studies of chronic diseases such as carcinoma and
tuberculosis
Reference range: Conventional unit Done in 2 readings after an hour and after 2 hours
Male: 47 vol.% Anticoagulant: 3.8% Na Citrate
Female: 43 vol.%
3. Brays Method
b) Haydens modification Anticoagulant: 1.3% Na Oxalate
Anticoagulant is 1.1% of NaC2O4 (Na Oxalate)
of 3.8% Na Citrate 4. Cutler Method
c) Van Allens modification method Anticoagulant: 3.8% Na Citrate
1.6% or 1.3% NaC2O4 or 3.8% Na Citrate
d) Sanford Magath 5. Micro Method
1.1% of 1.3% Na oxalate or 3.8% Na citrate Especially for neonates
e) Brays Method a) Landau Smith Method
Heparin is the anticoagulant b) Smith Method
2) Micro method
a) Micro Adams method Most popular and simple
Uses heparinize capillary tube
Mix to avoid coagulation CONDITIONS OF ESR
Seal with clay sealer then wax r FASTER RATE ESR
Spin: 10,00012,000 rounds per minute (RPM) for Pregnant women NON-PATHOLOGICAL
5minutes Menstruating women
Read with micro hematocrit capillary reader Suffering with:
Tuberculosis PA
TH
Cancer OL
OG
Rheumatic fever IC
AL
Malignant lymphoma
!!!!!!0$

r SLOWER RATE ESR

Spherocytosis
!

Poikilocytosis PA
!

TH
Sickle cell anemia OL
OG
Severe Iron Deficiency Anemia (IDA)
!

I CA
Thalassemia L
!

Jaundice
ESR$Scale$100$!!!!

Note:
The first drop of blood is usually discarded because it is
contaminated with dead epidermal cells and tissue juices.

 TRANSCRIBED GEN CAMATO

HEMATOLOGY LABORATORY
HEMATOLOGY 1 | LABORATORY

ZETA SEDIMENTATION RATIO

Determines the suspension stability of RBC that is not
affected by anemia
Measures the degree of packing of erythrocyte that occurs
during a 45 degrees cycle of dispension and compaction of
RBC in a special capillary tube and special centrifuge:
zetafuge
Zetafuge four spin cycle of 400 RPM/45 sec. For 4 mins.
Reference Value: 4051%

HEMOCYTOMETRY
Numerical evaluation of the formed elements of blood
Estimation of the number of blood cells in a given
volume of blood

METHODS FOR HEMOCYTOMETRY * OBSOLETE METHODS; NO LONGER USED

1) Turbidimetry
Base on the assumption that the more turbid the solution,
the more cells are present.

2) Microscopic Method * MANUAL METHOD

Using pipet method/techniques or test tube dilution
method
Principle: blood cells are counted under the microscope
with the use of counting chamber, pipets, and diluting fluid 2. Fuchs Rosenthal
Design for CSF analysis
TYPES OF COUNTING CHAMBER Depth: 0.2 mm
1. Spencer New Improved Neubauer Counting Chamber
Open type 3. Speirs Levy
It has 2 center flat form Has 4 center flatforms
1 ruled are divided in 9 primary square (1 sq.mm)
4 large square use for count PIPETS (Automatic and Non-Automatic)
1) Unopette * COMMONLY USED; PREFERRED FOR PLATELET COUNTING
Central primary square count [ Microblast capillary pipet that automatically suck just
Depth: 0.1 mm
the right amount of a sample, connected into a plastic
container, containing just the right amount of diluting
fluid

2) Thoma pipet *MANUAL

CLEANING OF THE PIPETS

1. Wash with water, alcohol and then ether. Air dry
2. Wash with water, acetone. Air dry

RBC Pipette WBC Pipette

Color of Bead
Size of Bulb Larger Smaller
Calibration Mark 101 11
Size of Lumen Smaller Larger

 TRANSCRIBED GEN CAMATO

HEMATOLOGY LABORATORY
HEMATOLOGY 1 | LABORATORY

STEPS IN HEMOCYTOMETER (Continuation RBC dilution)

A. Pipet Method ! Formol citrate (Dacies fluid)
1. Suck blood up to 0.5 mark of the pipet Best RBC diluting fluid
2. Suck diluting fluid up to 101 (for RBC) and 11 (for WBC) With preservative action so NO MOLD formation
3. Shake pipet to mix Cell morphology not altered
4. Discard first few drops
5. Charge counting chamber ! NSS
6. Count RBC in 5 intermediate square and WBC in 4 corner Used in cases of emergency
larger square Ideal to use in case of excessive rouleaux formation or
agglutination.
B. Test tube dilution Method
1. Add 4mL of dilution fluid
2. Shake tube
3. Obtain sample by a capillary tube WBC DILUTING FLUID * Function: to LYSE RBC
4. Charger counting chamber ! 1-3% Acetic acid with Gentian violet
5. Count in counting chamber ! 1% HCl
! Tuerks
Note: Acetic acid
to convert mL to L Methyl violet
ex. 0.02 mL x 1000 L = 20 L Distilled water
1mL

2 TYPES OF AUTOMATED HEMOCYTOMETRY

1) OPTICAL AUTOMATED
COMPUTATION FOR Principle: blood cells are counted as deflections of light
3
RBC in Millions/mm = RBC count x 10 x 200 x 5 beams passes to the dark field area of the machine.
Example: Fisher autocytometer
I Wherein: 10 depth correction factor (constant)
200 dilution factor (variable)
5 area correction factor (constant) 2) ELECTRICAL AUTOCYTOMETER
Principle: blood cells are counted as changes in voltage
COMPUTATION FOR pulse
3
WBC in Thousand/mm = WBC counted x 10 x 20 Example: Coulter counter (diluting fluid: Isoton)
4

I Wherein: 10 depth correction factor (constant) Inclusion Composed of Stain Indications

20 dilution factor (variable)
Disturbed
4 area correction factor (constant) erythropoietin
Hemolytic
CHARACTERISTIC OF IDEAL DILUTING FLUID Howell-jolly DNA Wright
Anemia
Isotonic (RBC) Hypotonic (WBC) Megaloblastic
9 0.85 -0.9% NaCl 9 to Lyse RBC anemia
* To prevent RBC from Shrinking & Swelling Post-
Chead and economical splenectomy
Easy to secure and prepare Wright
Basophilic Thalassemia
With preservative action RNA New Methylene
stippling Lead poisoning
Blue
With high specific gravity
Stable Pappenherimer G6PD deficiency
Denatured
With buffer action bodies
precipitated Supravital Stain
Thalassemia
Non-allergenic Siderotic Unstable
hemoglobin
granules hemoglobins
Non-corrosive
Remnants of Megaloblastic
RBC DILUTING FLUID Cabots ring Wright
mitotic spindle anemia
! Hayems
Initiates mold and rouleaux formations
Malaria
Can stand for a long period of time and no corrosive Parasites Babesia Wright
Parasitic
effect infection
Trypanosomes

! Gowers
Prevents rouleaux formation and precipitation of protein

! Toissons
Initiates mold formation so it should be filtered
With high specific gravity
With stain so used by beginners since RBC are easily

identified form WBCs whose nuclei stain are blue. A wise man will hear, and will increase learning; and a man of understanding shall
attain unto wise counsels:
- Proverbs 1:5