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Week 8

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8 views

Week 8

Uploaded by

gomedina1013lag
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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1.

Understand the principle of Osmotic Fragility test


2. Know the proper procedure of performing Osmotic fragility test
3. Interpret the result properly
4. Know the conditions associated with an increased or decreased
Osmotic fragility test result
The student/s will become familiar in the proper procedure of
osmotic fragility test. The students will answer the guide questions
after the experimentation.

The osmotic fragility test is a measure of the ability of red cells to take
up fluid without lysing. This procedure is employed to help diagnose
different types of anemia, in which the physical properties of red cells
are altered.
Red cells suspended in hypotonic solution of sodium chloride take up
water, swell, become spheroidal and after reaching the critical volume
eventually burst. The cell that is thicker than normal (spherocyte in
Hereditary Spherocytosis) has a decrease surface/volume ratio, and its
capacity to expand is limited.

Consequently, it burst upon intake of small amount of water in


relatively high concentration of the salt. Its osmotic fragility is
increased and its osmotic resistance is decreased
On the other hand, the thin or flat cells in hypochromic anemia can
take up considerable amounts of water (increased surface/volume
ratio) and reaches the critical volume of solution that has a lower
concentration of dissolved particles as to that solution with which it
is compound.
Materials:
 12 Kahn tubes or ordinary test tubes
 Test tube rack
 0.5% NaCl
 Distilled water
 Capillary pipette or Sahli pipette
 Anticoagulated blood (Heparinized blood)
TEST TUBE RACK
KHAN TUBE
SAHLI PIPETTE HEPARINIZED BLOOD
1. Arrange 12 tubes in a rack. Label it from number 25 up to number 14, starting from the
left.
2. With the capillary or Sahli pipette, place in each tube the number of drops of stock 0.5%
NaCl indicated by the number of the test tube.
Note: Hold the pipette at the same angle to ensure uniformity of drops
3. With the same pipette, add into each tube the number of drops of distilled water
required to bring the value in each tube to 25 drops.
TEST TUBE NUMBER 25 24 23 22 21 20 19 18 17 16 15 14
Drops of 0.5% NaCl 25 24 23 22 21 20 19 18 17 16 15 14
Drops of Distilled Water 0 1 2 3 4 5 6 7 8 9 10 11
% NaCl 0.50% 0.48% 0.46% 0.44% 0.42% 0.40% 0.38% 0.36% 0.34% 0.32% 0.30% 0.28%
(Concentration of NaCl)
4. Mix by lateral tapping
5. Draw blood into a Sahli-Hellige pipette up to 20 ul or 0.02 mL and dispense in each test
tube
TEST TUBE NUMBER 25 24 23 22 21 20 19 18 17 16 15 14
Drops of 0.5% NaCl 25 24 23 22 21 20 19 18 17 16 15 14
Drops of Distilled Water 0 1 2 3 4 5 6 7 8 9 10 11
% NaCl 0.50 0.48 0.46 0.44 0.42 0.40 0.38 0.36 0.34 0.32 0.30 0.28
(Concentration of NaCl)
Amount of blood Add 20 uL or 0.02 mL to all tubes

6. Mix by inverting the tube once or twice


7. Stand for two hours
 Check for initial and complete hemolysis
 Initial hemolysis: tinge of pinkness in the supernatant and some cells at the bottom of the tube
 Complete hemolysis: clear, red supernatant with no microscopic cells at the bottom of the tube
Hemolysis = Tube number with initial and complete hemolysis x 0.02 ml (amount of blood in each tube)

Initial hemolysis: Tube number 21 x 0.02 = 0.42%


Final hemolysis: Tube number 16 x 0.02 = 0.32%

Initial hemolysis: 0.42-0.44%


Complete hemolysis:0.32-0.34%

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