ATP Determination Kits

In energy consuming biological reactions the level of ATP is the essential indicator for enzyme activity or cell viability. The Biaffin ATP Determination Kits offer convenient bioluminescense assays for quantitative determination of small amounts of pre-existing ATP or ATP formed in enzymatic reactions. Catalysed by firefly luciferase the substrate D-luciferin is oxidized in an ATP-dependent process generating chemiluminescence at 560 nm (pH 7.8):

The timestable assay is optimized for high throughput screening with nearly constant luminescence signals over a period of up to four hours. The sensitive assay can detect nanomolar to micromolar concentrations of ATP.

The sensitive assay is optimized for fast determination of low ATP levels. After a 10 min incubation of the assay reagent, ATP concentrations down to 0.1 pmol can be exactly determined using the linear luminescent signal of the luciferase reaction. Loss of luminescent signal and sensitivity is observed after incubation times of more than 30 minutes (for applications where longer incubation times are indispensable use the timestabe assay instead).

Both assays are available in packing units of 10 ml for 200 – 1000 assays or 100 ml sufficient for 2000 – 10000 assays.

The Kits consist of four components, which can be shipped at room temperature:

• Firefly Luciferase (Component A, ready to use glycerol stock solution)
• D-Luciferin (Component B, solid, to dissolve in reaction buffer)
• Dithiothreitol DTT (Component C, solid, to dissolve in reaction buffer)
• Reaction Buffer (Component D)

The final reaction mix can easily be prepared and stored light protected in appropriate aliquots at –20oC. The standard reaction for both assays can be performed by adding an equal volume of the final assay mix to the ATP solution (usually 10 to 50 µl) in a 96 or 384 well plate optimised for luminescent reading. The luminescent signal can be measured preferably in a luminometer after 10 minutes at room temperature. As an alternative to a luminometer a scintillation counter can be used to measure luciferase activity. Unknown ATP concentrations can be exactly and reproducible determined using a standard ATP concentration curve generated under comparable experimental conditions (temperature, incubation times, assay volume, luminometer adjustments, etc.)

More detailed information about the assay conditions, prices and order placement at www.proteinkinase.de/html/luciferin.html

BIAFFIN GmbH & Co KG
Biomolecular Interaction Analyses
Heinrich-Plett-Str. 40
D-34109 Kassel
Germany
Tel.: +49 (0) 561 804 4661
Fax: +49 (0) 561 804 4662
Mail: info@biaffin.com
www.biaffin.com
www.proteinkinase.de