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Short answer: Nitro blue tetrazolium (NBT, also spelled nitroblue tetrazolium) is a tetrazolium salt, CAS 298-83-9, that turns blue when it is reduced. The same compound sits behind three different things people search for: a clinical blood test that screens for chronic granulomatous disease, a colour-development substrate paired with BCIP on blots and tissue sections, and a redox dye. Which one you need decides everything else.
This page is general reference for laboratory and clinic staff. It is not medical advice and does not interpret anyone’s test result.
Definition and identifiers
NBT is listed in PubChem (CID 9281) as nitroblue tetrazolium chloride. The identifiers below are those PubChem records, checked 2026-11-07.
| Property | Value |
|---|---|
| Name variants | Nitro blue tetrazolium, nitroblue tetrazolium, NBT, nitrotetrazolium blue chloride |
| CAS number | 298-83-9 |
| Molecular formula | C40H30Cl2N10O6 |
| Molecular weight | 817.6 |
What is the CAS number for nitro blue tetrazolium?
The CAS Registry Number for nitroblue tetrazolium chloride is 298-83-9, the same number vendors print on their nitrotetrazolium blue chloride specification sheets.
Which chemical is used in a tetrazolium test?
"Tetrazolium test" is a family name, not one reagent. Tetrazolium salts are a compound family used as colour-change reagents. NBT is the one used in the clinical nitroblue tetrazolium test. Other members of the family, such as MTT and XTT, are used in cell viability and proliferation readouts; see MTT assay, XTT assay and CCK-8 assay. If a protocol only says "tetrazolium", check the named salt before ordering or preparing anything.
What is the nitroblue tetrazolium test for?
According to MedlinePlus (review date 1 January 2025, checked 2026-11-07), the nitroblue tetrazolium test is a blood test used to screen for chronic granulomatous disease (CGD), an inherited condition. In the laboratory, NBT is added to the person’s white blood cells and the cells are examined under a microscope for a blue colour.
Nitroblue tetrazolium test positive or negative: what the wording means
MedlinePlus describes the expected finding as white blood cells that turn blue when NBT is added, and the opposite finding as no colour change. Because the colour change is the expected finding, a "positive" or "negative" label on a report can be read the wrong way round. Reference ranges vary by laboratory, so the reporting laboratory’s own wording governs. MedlinePlus lists no special preparation and the risks as those of any blood draw. Questions about a specific result belong with the ordering clinician.
How NBT is used at the bench
NBT/BCIP as an alkaline phosphatase substrate
In blots and immunohistochemistry, NBT is combined with BCIP as the chromogenic substrate for alkaline phosphatase-labelled detection. The manufacturer’s sheet for your product states the buffer and preparation requirements. Storage and stability differ between products and formats (tablet, powder, ready-made solution), so follow the sheet that came with your lot. For the surrounding workflow see how to interpret western blot results and immunohistochemistry controls and troubleshooting.
NBT as a redox dye
Because NBT changes colour when reduced, it is also used as a reduction indicator. Method details differ between assays, so use a published protocol for the specific measurement rather than a generic recipe. For the tetrazolium readouts used in cell work, compare the cell proliferation assay overview.
Three uses compared
| Use | Who runs it | What is observed | Source |
|---|---|---|---|
| NBT blood test | Clinical laboratory, ordered by a clinician | Blue colour in white blood cells examined under a microscope | MedlinePlus |
| NBT/BCIP blot or tissue stain | Research and pathology bench | Colour development at alkaline phosphatase-labelled sites | Manufacturer’s product sheet |
| Cell viability tetrazolium salts (MTT, XTT, CCK-8) | Cell culture labs | Colour from reduction by cells; different salts, not NBT | See the linked assay guides |
What is TTC chemical?
TTC is the abbreviation for 2,3,5-triphenyltetrazolium chloride. It is a different tetrazolium salt from NBT: same compound family, different molecule. Do not substitute one for the other because both are called tetrazolium.
Common misunderstandings
- One reagent, one test. NBT is used for the blood test and for the blot substrate, but the protocols, buffers and readouts have nothing in common.
- Two spellings mean two chemicals. Nitro blue and nitroblue are two spellings of the same compound.
- All tetrazolium salts are interchangeable. NBT, MTT, XTT and TTC are separate compounds; an assay is validated for the salt it names.
- Any buffer works with NBT/BCIP. Buffer requirements are product-specific, so check the buffer named on your product sheet.
Frequently asked questions
What is nitro blue tetrazolium used for?
Three things: the clinical NBT test that screens for CGD, the NBT/BCIP alkaline phosphatase substrate in blots and tissue staining, and as a tetrazolium redox indicator.
What is the molecular weight of nitroblue tetrazolium?
817.6, for the chloride salt with formula C40H30Cl2N10O6 (PubChem CID 9281).
How should NBT be stored?
Storage depends on the product format. Follow the storage temperature and light-protection instructions on the sheet for the exact product and lot you hold.
Sources
- MedlinePlus, Nitroblue tetrazolium test, review date 1 January 2025, checked 2026-11-07.
- PubChem, Nitroblue Tetrazolium Chloride, CID 9281, checked 2026-11-07.
