blood exercises (phyana lab)
TRANSCRIPT
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PHY-ANA LABBLOOD
EXERCISES
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COMPLETE BLOOD COUNT
Hematocrit
Hemoglobin Differential white blood cell
Red blood cell
White blood cell
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HEMATOCRIT
% of formed cells in whole blood
99% RBCs and 1% WBCs and platelets
Estimate if RBCs are adequate
Greek hematoblood andcrit to
judge
Aka packed cell volume (PCV), Hctor erythrocyte volume fraction (EVF)
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HEMATOCRIT
One of the simplest, most accurate, & valuable tests
Detecting cases of anemia
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HEMATOCRIT
Specimen
Fresh capillary blood (with heparin)
Adams Microhematocrit Method
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HEMATOCRIT
1. Blood of thecapillary tube
2. Sealing clay (3 mm)
3. Centrifuge 10,000rpm for 4-5 minutes
4. Level of packed RBCusingmicrohematocritreader
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HEMATOCRIT
Capillary tube withseal towards the
outside
Balance tubes inthe centrifuge
Securely screw thecover of thecentrifuge
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HEMATOCRIT
When rotation has stopped,
remove tube Take note of the appearance
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HEMATOCRIT
Capillary tubewith seal
toward thecenter
Align upper
portion of theseal with theblack line
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HEMATOCRIT
Rotate thewhole assembly
so that the pinstops (100mark)
Rotate theupper disk tomove the curve
line with the top
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HEMATOCRIT
Rotate the entire assemblyuntil the curved line is lined
up with the boundarybetween packed RBC andplasma
Read the % packed cells atthe right
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HEMATOCRIT
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HEMOGLOBIN
Red-pigmented protein
Transports oxygen andcarbon dioxide
Measured as
oxyhemoglobin Indirectly measured byconverting to compounds
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HEMOGLOBIN
Acid Hematin
Method Yellowish brownsolution is comparedto the color standard
in the comparatorblock
Darker the color =higher Hgb content
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HEMOGLOBIN
1. 0.1N HCl2 mark of Sahlis tube
2. Aspirate 0.02 mL blood using Sahlis pipette
3. Expel the blood sample to the tube4. Rinse the pipette with dist. water 3xadd to the
mixtureStand 10 mins
5. Add dist. water drop by drop (mix with stirring rod)
until color matches with the block6. Readinglower meniscus
7. Report gm% or gm/dL or gm/100mL (CU) and ingm/L (SI)
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DIFFERENTIAL WBC COUNT
Examination of a thin smear determining the
percentages of WBC types
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DIFFERENTIAL WBC COUNT
Specimen
EDTA blood
Within 2-3 hours ofcollection
Within the mark of thetube
Avoid
Old specimen
Excessive amount of
anticoagulant to specimen
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DIFFERENTIAL WBC COUNT
Blood smear
preparation Most important step
Two-Slide or Wedge
Method Simplest
Most popular
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DIFFERENTIAL WBC COUNT
1. Drop of blood from mixed
sample on a clean glass slide2. Spreader slide at an angle of
about 30-45o
3. Allow blood to spread evenly;Control thickness of smear
4. Air dry
Do not blow dry: RBC artifact
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DIFFERENTIAL WBC COUNT
Good smear:
Thick to thin Occupy 2/3 or
Smooth and evensurface
Free from ridges,waves, holes
Margin-free
Feathery edge
Factors that affect:
Angle of the spreaderslide
Greater angle: thicker &shorter
Size of the blood drop Speed of spreading
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DIFFERENTIAL WBC COUNT
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DIFFERENTIAL WBC COUNT
Staining of Blood Smears
Methanol: fixative (30s) Eosin: acidic dye (6s)
Stains Hgb & leukocytes
Methylene blue: basic dye
(4s) Nucleoproteins, nucleic acids
Buffer solution (pH 7.2) for
45s
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DIFFERENTIAL WBC COUNT
Staining of BloodSmears
Dip Method Rapid)
Quick method
Modified Wright-Giemsa
buffered in methanol atpH 6.8
Tightly sealed
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DIFFERENTIAL WBC COUNT
Blood Smears
RBC: pink to salmon Nucleus: dark blue to purple
Neutrophils: lavender to lilac
Basophils: dark blue to black Eosinophils: red to orange
Area between cells: colorless,clean and free of precipitates
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DIFFERENTIAL WBC COUNT
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DIFFERENTIAL WBC COUNT
Smear Examination
1. LPO Assess overall quality Rapid detection of large
abnormal cells
Not overlapping or too scanty2. Shift to OIO
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DIFFERENTIAL WBC COUNT
Method of Differential Counting
Battlement Count 100 white blood cells whiledifferentiating
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DIFFERENTIAL WBC COUNT
Method of Differential Counting
Battlement Count 100 white blood cells whiledifferentiating
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DIFFERENTIAL WBC COUNT
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HEMACYTOMETER
Counting chamber
WBC pipette RBC pipette
Accessory devices
Suction device Thick cover slip
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COUNTING CHAMBER
Heavy, colorless glass
3 parallel platformsseparated by moats
Central: 0.1 mm lowerthan lateral
Transverse groove
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COUNTING CHAMBER
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COUNTING CHAMBER
1 Primary square
3x3 mm (9 sq. mm)
9 Secondarysquares
1x1 mm
4 corners: WBCcount
16 tertiary squares
W1, W2, W3, W4 64 squares
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COUNTING CHAMBER
Central secondarysquare
25 tertiary squares 0.2 mm each
16 quaternarysquares
Total number ofquaternary: 400
RBC count
5 tertiary squares:80 squares
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DILUTED BLOOD
PREPARATION 0.5 mark: blood
Diluting fluid
11 WBC, 101RBC
Constant
rotation Over aspirate
or presence of
bubbles: repeat
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CHARGING
Cover slip
No dirt, thumb marks,
tissue strands Discard
WBC 2-3 drops
RBC 5-6 drops Angle of the pipette
(30-35)
Stand for 5-10 minutes
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RBC AND WBC THOMA
PIPETTES RBC pipette
Stem 0.0 to 1.0 contains1 unit of volume
Mixing chamber or bulb0.1 to 101 holds 100units of volume
WBC pipette
Stem 0.0 to 1.0
Bulb 1.0 to 11
Stem volume is 10x the
bulb volume: 10 units
RBC
PIPETTE
WBC
PIPETTEUpper mark 101 11
Bore Smaller Bigger
Bead Red White
Dilution 1:200 1:20
Size of bulb Bigger Smaller
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STANDARD PATTERN OF
COUNTING Cells touching
any of the lines
on the top andleft borders areincluded
Cell difference
between 2squares
RBC 20 or less
WBC 12 or less
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COUNTING CELLS
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COMPUTATION
RBC COUNT No. of cells/cumm = total number of cells counted
area X depth X dilution
= total number of cells counted
1/5 X 1/10 X 1/200
= cells counted X 10,000
Normal values: male: 4.5-6.0 M/cumm
female: 4.0-5.5 M/cumm
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COMPUTATION
WBC COUNT
No. of cells/cumm = total number of cells counted
area X depth X dilution
= total number of cells counted
4 X 1/10 X 1/20
= cells counted X 50
Normal value: 5,000-10,000/cumm
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BLOOD GROUPS
Antigens
Antibodies
Agglutination
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ABO BLOOD GROUPING
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BLEEDING TIME
ukes Method
Finger prick
Allow blood to flow freely
Start time: drop of bloodappears
Blot with filter paper Do not touch the wound
Stop time: when bleeding stops
Normal: 1-3 minutes
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COAGULATION TIME
Drop or Slide Method
Prick
Drop of blood on a slide
Start: when in contact with the slide
Tip of lancet every 30 second
interval Observe fibrin formation
Stop timer
Normal: 3-6 minutes
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COAGULATION TIME
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HYPEREMIA OR
CONGESTION Note the skin color, blood vessel
condition, temperature of left indexfinger
Immerse in hot water (60C) for 5minutes
Note the changes and thesensation felt
Rubber band (5 minutes)2ndinterphalangeal joint
Note the changes and thesensation felt
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HYPEREMIA OR
CONGESTION Hyperemia: active increase in
blood volume
Dilation
Physiological: blushing or duringexercise
Reddish
Congestion: passive increase in
volume of blood Impaired venous blood flow orvenous obstruction
Reddish-blue (cyanosis)
Always pathological
Cardiac failure
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CAPILLARY RESISTANCE
TEST Assesses the fragility
of capillary walls
Hemorrhagictendency
Thrombocytopenicpurpura
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CAPILLARY RESISTANCE
TEST Thrombotic Thrombocytopenic Purpura
(clots)-(low platelet number)-(purple bruises)
Rare blood disorder
Blood clots form in capillaries
Uses up platelets Bleeding problems
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CAPILLARY RESISTANCE
TEST Tourniquet Test Rumpel-Leede or Hess)
Mark red spots on the arm
Wrap the cuff of sphygmomanometer around
Inflate to 100 mmHg (5 mins) or 50 mmHg (10mins)
Release pressure (15-20 mins elapse)
Count the number of petechiae (ventral)
Interpret results
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CAPILLARY RESISTANCE
TEST Interpretation of results
Number of petechiae Grade
0-10 1+
11-20 2+
21-50 3+
51 and above 4+