Principle
This test detect the presence of HbS depend on the decreased solubility of this haemoglobin at low oxygen tension. In deoxygenated state, insoluble crystals are formed due to polymerization of HbS. This insoluble crystal leads to the characteristic shape change of RBCs called sickle cells.
Primary sample
2 ml whole blood in EDTA vaccute
Equipment and reagents
1) Sodium metabisulphite powder
2) Test tube & micropipette
3) Distilled water
Steps of procedure
1) Prepare 2% sodium metabisulphite (50 mg sodium metabisulphite powder in 2.5 ml distilled water)
2) Add 5 drops of the freshly prepared reagent to 1 drop of anticoagulated blood on a slide.
3) Seal between slide and cover glass with a petroleum jelly / paraffin wax mixture or with nail varnish.
4) Observe for sickle cells at the end of ½, 1 hr, 4 hr & 24 hours.
Result
Sickle cells are elongated cells with one or both pointed ends. They are boat-shaped, elliptical sickle or typical sickle-shaped.
Interpretation
Sickling takes place almost immediately in sickle cell anemia. Sickling should be obvious in sickle cell trait.
Interference
False positive:
1) Unstable hemoglobin
False negative:
1) Very low hemoglobin
2) Outdated reagent
3) Infants less than 6 months
4) HbS level less than 20%
False positive or false negative result can be minimised by using packed cells instead of whole blood.
Control
A test on positive control of HbA and HbS must be performed at the same time.