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Catalog # HK328

TCC, Human, ELISA kit

685,001.108,00

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The three distinct activation pathways of complement converge with the formation of a C5 convertase and it is cleavage of C5 by these convertases that initiates the lytic or terminal pathway. In contrast to the activation pathways, which require enzymatic cleavage for activation, the terminal pathway relies on conformational changes induced by binding. Binding of C6 facilitates binding of C7 which alters the conformation of the complex. After binding of C8 a variable number of C9 molecules associate with the C5b678 complex, the so called terminal complement complex (TCC) or Membrane Attack Complex (MAC). The formation of the membrane form of TCC causes lysis of cells or can trigger a variety of cellular metabolic pathways resulting in the synthesis and release of inflammatory mediators. The TCC contains neoantigens that are absent from the individual native components. Neoantigens are present both in the membrane-bound (MAC) and the fluid-phase (SC5b-9) complex. TCC is present in normal human plasma, <1 AU/ml, and increases in patients with complement activation.

Application
The human terminal complement complex (TCC) ELISA kit is to be used for the in vitro quantitative determination of human TCC in plasma, urine, cerebrospinal fluid, joint fluid and cell culture supernatant samples.

Principle
The human TCC ELISA is a ready-to-use solid-phase enzyme-linked immunosorbent assay based on the sandwich principle with a working time of 3½ hours. The efficient format of 2 plates with twelve disposable 8-well strips allows free choice of batch size for the assay. Samples and standards are captured by a solid bound specific antibody. Biotinylated tracer antibody will bind to captured TCC. Streptavidin-peroxidase conjugate will bind to the biotinylated tracer antibody. Streptavidin-peroxidase conjugate will react with the substrate, tetramethylbenzidine (TMB). The enzyme reaction is stopped by the addition of oxalic acid. The absorbance at 450 nm is measured with a spectrophotometer. A standard curve is obtained by plotting the absorbance (linear) versus the corresponding concentrations of the TCC standards (log). The TCC concentration of samples, which are run concurrently with the standards, can be determined from the standard curve.

Linearity
The linearity of the assay was determined by serially diluting a sample containing 250 mAU/ml human TCC. The diluted samples were measured in the assay. The line obtained a slope of 0.967 and a correlation coefficient of 0.999.

Recovery
Normal human blood samples (plasma), containing baseline levels of human TCC, were spiked with human TCC in concentrations of 2500 and 250 mAU/ml. Samples with and without TCC were incubated for 1 hour at 4° C. Samples were measured by ELISA. Values for TCC ranged between 153% and 128% (mean 140%).

Product type
Assays
Quantity
1 x 96 det., 2 x 96 det.
Standard range
8.2 to 2000 mAU/ml
Detection level
8.2 mAU/ml
Working volume
100 µl/well
Species
Human
Cross reactivity
Rabbit – Yes
Storage and stability
Product should be stored at 4 °C. Under recommended storage conditions, product is stable for at least six months.
Precautions
For research use only. Not for use in or on humans or animals or for diagnostics. It is the responsibility of the user to comply with all local/state and Federal rules in the use of this product. Hycult Biotech is not responsible for any patent infringements that might result with the use of or derivation of this product.
Disease
Infectious diseases, Nephrology
Safety Data Sheet Assay Manual
HK328
460 kb

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