MTT cell proliferation assay kit, 500 Rxn

MTT cell proliferation assay kit, 500 reactions. Kit Content: Solution A 5ml, Solution B 50ml. MTT Proliferation Assay Kit provides an easy to use tool for studying the induction and inhibition of cell proliferation in any in vitro model. This kit will also allow investigators to screen drug candidates involved in cell cycle regulation. In this assay, MTT is taken up by cells through the plasma membrane potential and then reduced to formazan by intracellular NAD(P)H-oxidoreductases.

MTT Cell Proliferation Assay Kit

Catalog Number:

CP-001, 1000 reactions

CP-002, 500 reactions

Introduction

The reduction of tetrazolium salts is now recognized as a safe, accurate alternative to radiometric testing. MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay, first described by Mosmann in 1983, is based on the ability of a mitochondrial dehydrogenase enzyme from viable cells to cleave the tetrazolium rings of the pale yellow MTT and form a dark blue formazan crystals which is largely impermeable to cell membranes, thus resulting in its accumulation within healthy cells. Solubilization of the cells by the addition of a detergent results in the liberation of the crystals which are solubilized. The number of surviving cells is directly proportional to the level of the formazan product created. The color can then be quantified using a simple colorimetric assay. The results can be read on a multi-well scanning spectrophotometer (ELISA reader).

Storage:

Upon receiving, Solution A MTT solution should be kept at-20⁰C and protected from light. Store properly, the kit components should remain stable for 6 months.

Kit Contents:

1.Solution A MTT solution: 5ml ( 500 reactions), 10ml (1000 reactions)

2. Solution B Crystal dissolving solutions: 50ml ( 500 reactions), 100ml (1000 reactions)

Protocol

1. Plate cells in 96-well tissue culture plate at density of 2x10⁵ per well in 100ul of culture medium.

2. 5 hours before the end of the incubation add 10ul of MTT solution from step one to each well containing cells.

3. Incubate the plate at 37ºC for 5 hours.

4. Remove media with needle and syringe.

5. Add 100ul of Solution B to each well and pipette up and down to dissolve crystals.

6. Transfer to plate reader and measure absorbance at 570nm.

Reference
Mosmann T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods. 1983 Dec 16;65(1-2):55-63.