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OUR SUPPLIERS

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aCella™ - TOX
Bioluminescence Non Radioactive Cytotoxicity Assay (GAPDH)
Key Benefits:
  • Safe - Non Radioactive Enzyme release assay.
  • Versatile - Useful for measuring activity of T Cells, Primary Cells, NK, complement and other lytic agents. Assay can be run in serum supplemented media.
  • Homogenous - One-step, no wash assay. Assay can be run in same plate as samples.
  • FAST - Results in 3-5 minutes. Chromium 51 or europium release for measurement are time consuming. The inherent sensitivity of luciferase detection is enhanced by the amplification effect of enzyme turnover, which produces thousands, millions or billions of high - energy  molecules for each molecule of the enzyme.
  • Highly Sensitive - Can detect fewer than 500 cells/well in the presence of serum or as few as 10 cells/well in serum-free or heat-killed media.
  • GAPDH: The fact that GAPDH is a natural component of cells, and does not need to be introduced into the cells in any manner, distinguishes this assay from all methods which require prelabelling of cells, transfection, transformation, or other methods of introducing proteins or other molecules into the target cells in order to generate a signal in a later step.
  • Advantages for measurement of cell mediated or complement mediated cytolysis - It is usually desirable to use smaller numbers of TCells than are needed for the 51 Cr – release assay, since excessive numbers of effector cells can increase the background signal. This is now possible due to the high sensitivity of aCella-Tox.
  • ADCC / CMC Assays - A non radioactive alternative to Cr 51 assays. Sample Protocol published.
  • HTS - Adaptable for High Throughput format
  • Non-destructive assay allows monitoring of additional parameters.
Introduction to aCella-TOX:
Cell Technology introduces aCella-TOX, a new and highly sensitive assay using our patented Coupled Luminescent technology for the detection of cytotoxicity (1). This assay quantitatively measures the release of Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH) from Primary Cells, mammalian cell lines, bacterial cells(1,2,3).

Other enzyme release assays(5,6,7) for example the Lactate Dehydrogenase (LDH) release assay (5,6,7), are inconvenient and/or slow and may suffer from low sensitivity as a result of the poor signal and interference by serum or phenol red present in the media. The ATP-release assay (8) is inconvenient and much less sensitive than aCella-TOX, and is unsuitable for use in a cytotoxicity assay because the lytic signal is indirect.

aCella-TOX can work in both these media formulations and allows overnight assays while retaining its sensitivity. The sensitivity of aCella-TOX is also greatly enhanced by the coupled luminescent signal-amplification system, which yields a strong luminescent signal from even small amounts of released enzyme.

Assay Principle:
GAPDH is an important enzyme in the glycolysis and gluconeogenesis pathways. This homotetrameric enzyme catalyzes the oxidative phosphorylation of D-glyceraldehyde-3-phosphate to 1,3-diphosphoglycerate in the presence of cofactor and inorganic phosphate.

In the aCella-TOX reaction scheme the release of GAPDH is coupled to the activity of the enzyme 3-Phosphoglyceric Phosphokinase (PGK) to produce ATP. ATP is detected via the luciferase, luciferin Bioluminescence methodology. Further, aCella-TOX is a homogeneous cytotoxicity assay; alternatively in dual mode, aCella-TOX can measure cytotoxicity and cell viability in the same plate. Culture supernatants can also be removed from the original plate and assayed in a different plate, allowing kinetics runs to be set up. The assay is non-destructive, allowing the monitoring of additional parameters such as gene expression.
 

Applications:
The aCella-TOX method has been tested with many modes of cytolysis, including;
  •  cellular cytotoxicity (T cells)
  •  complement (2,3), pore-forming agents,
  •  antibiotic-mediated lysis of bacteria, and
  •  detergent mediated and mechanical lysis
The method is highly general, since all known cells express copious amounts of GAPDH, and, unlike other enzymes, GAPDH is very readily released from the cytoplasm upon cell lysis. Using specially adapted formulations, the sensitivity of the method can be driven below 1 eukaryotic cell (2), which is impossible with any other reported liquid-phase method. Please consult with us if you have an application requiring specialized techniques.
Use of aCella-TOX for Measurement of Cell-Mediated (T Cells, ADCC, NK) or Complement-Mediated Cytolysis
The following kits are available:
Catalog #
 
Size* Price
CLATOX 100-3
 
500 €595
CLATOX 100-4 1000 €1145