Assignment task:
NBT is a yellowish, soluble compound that when reduced is converted to Formazan, a blue insoluble compound.
NADPH Oxidase is activated in neutrophils during the oxidative burst response. The oxygen radicals formed during these responses can reduce NBT. Therefore, the presence of the blue, insoluble Formazan deposits indicates a working oxidative burst mechanism.
Normally NADPH Oxidase is only activated when neutrophils detect an infectious microorganism (such as when their TLR receptors bind a bacterial ligand). In the lab, rather than use a whole infectious organism, a variety of mechanisms exist for stimulating neutrophils to activate NADPH Oxidase (including using solutions of latex toxin or bacterial filtrate [pieces]).
Brief Procedure: Whole blood is collected from the patient and some blood is combined with NBT solution. The blood-NBT mixture is mixed and incubated for about 10 minutes. The mixture is then smeared on a slide and cells are stained for identification. The appearance of cells is observed; the numbers of neutrophils with and without formazan deposits are counted.
Both a stimulated and unstimulated NBT test can be performed. In the unstimulated test the procedure above is followed. In the stimulated test both NBT and a stimulation solution (such as a solution containing latex toxin) are combined with the blood. Incubation, smearing, staining, and counting of cells is the same in both the stimulated and unstimulated tests.
If you were to perform the stimulated and unstimulated NBT test on normal individuals what would you expect to see?
If you were to perform the stimulated and unstimulated NBT test on individuals whose oxidative burst responses did not work what would you expect to see?