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CT01-5

Sigma-Aldrich

MTT Reagent A

Useful in the study of cell & mitochondrial health, MTT is a pale yellow substrate that is cleaved by living cells to yield a dark blue formazan product.

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Synonym(s):
MTT Formazan Assay
eCl@ss:
32161000
NACRES:
NA.32

Quality Level

form

solid

manufacturer/tradename

Chemicon®

technique(s)

activity assay: suitable
cell based assay: suitable

shipped in

wet ice

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This Item
CT01CT02475989
Sigma-Aldrich

CT01-5

MTT Reagent A

Sigma-Aldrich

CT01

MTT Cell Growth Assay Kit

Sigma-Aldrich

CT02

MTT Cell Growth Assay Kit

MTT Membrane-permeable yellow dye that is reduced by mitochondrial reductases in living cells to form the dark blue product, MTT-formazan.

475989

MTT

manufacturer/tradename

Chemicon®

manufacturer/tradename

Chemicon®

manufacturer/tradename

Chemicon®

manufacturer/tradename

Calbiochem®

technique(s)

activity assay: suitable

technique(s)

activity assay: suitable, cell based assay: suitable

technique(s)

activity assay: suitable, cell based assay: suitable

technique(s)

-

shipped in

wet ice

shipped in

wet ice

shipped in

wet ice

shipped in

ambient

Quality Level

100

Quality Level

100

Quality Level

100

Quality Level

100

General description

Reagent A: MTT, (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide), 50mg/vial.

MTT is a pale yellow substrate that is cleaved by living cells to yield a dark blue formazan product. This process requires active mitochondria, and even freshly dead cells do not cleave significant amounts of MTT. Reagent A is a component of Chemicon′s MTT assay kit, Catalog Number CT01/CT02.

Application



Procedure:
1. Carry out a lymphokine, mitogen, or complement-mediated cytotoxicity assay using standard methods, in 96-well flat-bottomed tissue culture plates of good optical quality (e.g. Falcon). The final volume of tissue culture medium in each well should be 0.1 mL, and the medium (e.g. RPMI or DMEM) may contain up to 10% Fetal Bovine Serum.

2. At the end of the assay add 0.01 mL AB Solution (MTT) to each well. Mix by tapping gently on the side of the tray.

3. Incubate at 37°C for cleavage of MTT to occur. Optimal times may vary according to the assay, but four hours is suitable for most purposes. At the end of this time, the MTT formazan produced in wells containing live cells will appear as black, fuzzy crystals on the bottom of the well.

4. Add 0.1 mL Solution C to each well. Mix thoroughly by repeated pipetting with a multichannel pipettor. The HCl converts the phenol red in tissue culture medium to a yellow color that does not interfere with MTT formazan measurement. The isopropanol dissolves the formazan to give a homogeneous blue solution suitable for absorbance measurement.

5. Within an hour, measure the absorbance on an ELISA plate reader with a test wavelength of 570 nm and a reference wavelength of 630 nm. After several hours at room temperature, serum proteins may begin to precipitate due to the acid/alcohol. Chilling the plates will hasten the precipitation. If the plates must be stored before measuring, keep at 4° C before adding the acid/alcohol, then warm to room temperature and add acid/alcohol just before reading.

Results:
The MTT assay will normally detect 200 to 50,000 cells of a typical cell line, although 1,000 to 50,000 is the useful range. This number may vary for other cell types. Cytotoxic assays should be set up so that the control, unlysed cells give a signal of 0.2 to 0.4, and proliferation assays should yield a similar value at plateau concentrations. This corresponds to about 20-50,000 cells per well with a typical cell line.

Absorbance is directly proportional to the number of cells; actual cells do not absorb significantly, even up to concentrations of l x 106 cells/mL.

Legal Information

CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany

pictograms

Exclamation markHealth hazard

signalword

Warning

Hazard Classifications

Eye Irrit. 2 - Muta. 2 - Skin Irrit. 2 - STOT SE 3

target_organs

Respiratory system

Storage Class

11 - Combustible Solids

wgk_germany

WGK 3

flash_point_f

Not applicable

flash_point_c

Not applicable


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Rapid colorimetric assay for cell viability: application to the quantitation of cytotoxic and growth inhibitory lymphokines.
Green, L M, et al.
Journal of Immunological Methods, 70, 257-268 (1984)
Zeynep Firtina Karagonlar et al.
Cells, 9(5) (2020-05-16)
The complex and heterogeneous nature of hepatocellular carcinoma (HCC) hampers the identification of effective therapeutic strategies. Cancer stem cells (CSCs) represent a fraction of cells within tumors with the ability to self-renew and differentiate, and thus significantly contribute to the

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