Cytotoxicity Assay Kit

Catalog No. ABIN1000305

Product Details

Specificity: 50 cells

Characteristics: Safe. Non-radioactive assay (cf. 3 H-thymidine incorporation assay).
Sensitive and accurate. As low as 50 cells can be quantified.
Homogeneous and convenient. Mix-incubate-measure type assay. No wash and reagent transfer steps are involved.
Robust and amenable to HTS: Z' factors of 0.6 to 0.7 are routinely observed in 96-well and 384-well plates. Can be readily automated on HTS liquid handling systems for processing thousands of samples per day.

Components: Assay Buffer: 10 mL. Substrate: 120 µL. ATP Enzyme: 120 µL.

Application Details

Application Notes: Cell proliferation: effects of cytokines, growth factor, nutrients.
Cytotoxicity and apoptosis: evaluation of toxic compounds, anti-cancer antibodies, toxins, environmental pollutants etc.
Drug Discovery: high-throughput screening for anticancer drugs.

Comment:

After adding the Reconstituted Reagent, the luminescence signal is stable for about 15 min and decreases slow thereafter. Reading is best performed within 30 min.

Protocol: 1. Cell Culture. Plate cells at 100 µL/well in white opaque tissue culture plates. If desired, add 5 µL test compounds and controls dissolved in PBS or culture medium per well. Rock plate lightly to mix and incubate for desired period of time (e.g. overnight).
2. Assay. Bring all components to room temperature. Keep thawed ATP Enzyme on ice or 4°C. For each test well, mix 95 µL Assay Buffer with 1 µL Substrate and 1 µL ATP Enzyme. Add 90 µL Reconstituted Reagent to each test well and mix by tapping the plate. Incubate for 2 minutes at room temperature. Read luminescence on a luminometer. For most luminometers (Berthold Luminometer, LJL Analyst, Top Count, MicroBeta Counters, CLIPR and LeadSeeker), integration time of 0.1 to 5 sec is appropriate. ASSAY

Restrictions: For Research Use only

Handling

Storage: -20 °C

References

Massa, Yang, Xie, Shi, Bilgen, Joyce, Nehama, Rajadas, Longo: "Small molecule BDNF mimetics activate TrkB signaling and prevent neuronal degeneration in rodents." in: The Journal of clinical investigation, Vol. 120, Issue 5, pp. 1774-85, (2010) (PubMed).

Target Details

Background: Rapid, bioluminescent luciferin/luciferase based assay for cell viability, proliferation, cytotoxcity, high-throµghput screening for anticancer agents.
Procedure: 2 min.

Adenosine 5'-triphosphate (ATP) is the chemical energy for cellular metabolism and is often referred to as energy currency of the cell. ATP is produced only in living cells during photosynthesis and cellular respiration and consumed in cellular processes including biosynthetic reactions, motility and cell division. It is a key indicator of cellular activity and has been utilized as a measure of cell viability and cytotoxicity in research and drug discovery. This Assay Kit provides a rapid method to measure intracellular ATP, cell viability and cytotoxicity. The single working reagent lyses cells to release ATP, which, in the presence of luciferase, immediately reacts with the Substrate D-luciferin to produce light. The light intensity is a direct measure of intracellular ATP concentration and hence number of living cells. This non-radioactive, homogeneous cell-based assay can be conveniently performed in microplates. The reagent is compatible with all culture media and liquid handling systems for high-throughput screening applications in 96-well and 384-well plates.