Description: Ficolin-3, Human, ELISA kit

Ficolins are a group of proteins containing both a collagen-like domain and a fibrinogen-like domain. Three forms of Ficolin have been identified in humans: Ficolin-1 (M-ficolin), Ficolin-2 (L-ficolin) and Ficolin-3 (H-ficolin 3). Ficolin-3, also known as Hakata-antigen or H-ficolin, is composed by a collagen-like strand and three C-terminal recognition domains which bind to acetyl groups on microbial surfaces such as GlcNAc or GalNAc. Ficolin-3 is synthesized both in the liver, from where it is secreted into the blood circulation, and in the lung. Similar to MBL and Ficolin-2, Ficolin-3 relies on MBL-associated serine protease 2 (MASP-2) for activation of the complement system. After binding of Ficolin-3/MASP-2 complexes to microbial surfaces, MASP-2 sequentially cleaves C4 and C2, thereby generating the C3 convertase C4bC2b, which finally leads to opsonization and direct lysis of pathogens and recruitment of inflammatory cells. Ficolin-3 is present in serum at mean concentration of 15 g/ml, with only minor variations. Ficolin-3 was present in all sera from more than 150,000 individuals tested, except in some systemic lupus erythromatosis patients. Approximately 5% of systemic lupus erythromatosis patients were found to be Ficolin-3 negative, probably owing to the presence of anti-Ficolin-3 autoantibody. In 398 patients with other autoimmune diseases, Ficolin-3 was always present. In liver disease the serum levels decreased with increasing severity of cirrhosis. Ficolin-3 bound to a population of late apoptotic cells, while a strong and uniform binding to necrotic cells was observed. The binding properties differed from those of MBL and Ficolin-2. Ficolin-3 binding to late apoptotic cells resulted in a significant increase in adhesion/uptake by macrophages.

Specifications

Catalog number HK340-02
Product type Assays
Quantity 2 x 96 det.
Standard range 3-500 ng/ml
Detection level 3 ng/ml
Working volume 100 l/well
Species Human
Alias Hakata-antigen, H-ficolin
Application The human Ficolin-3 ELISA has been developed for the quantitative measurement of natural Ficolin-3 in plasma, and serum.
Principle The human Ficolin-3 ELISA is a ready-to-use solid-phase enzyme-linked immunosorbent assay based on the sandwich principle with a working time of 3 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 Ficolin-3. 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 Ficolin-3 standards (log). The human Ficolin-3 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 98.1 ng/ml human Ficolin-3. The diluted samples were measured in the assay. The line obtained a slope of 0.883 and a correlation coefficient of 1.0.
Recovery Normal human blood samples (plasma), containing baseline levels of 500 ng/ml were spiked with recombinant Ficolin-3 in concentrations of 6.5 and 100 ng/ml. Samples with and without Ficolin-3 were incubated for 1 hour at room temperature. Samples were measured using the ELISA. Values for Ficolin-3 ranged between 94 % and 115%.
Storage and Stability Product should be stored at 4 C. Under recommended storage conditions, product is stable for one year.
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.
References 1. Holmskov, U et al; Collectins and ficolins: humoral lectins of the innate immune defense. Annu Rev Immunol 2003, 21: 547
2. Kuraya, M et al; Specific binding of L-ficolin and H-ficolin to apoptotic cells leads to complement activation. Immunobiol 2005, 209: 689
3. Honore, C et al; The innate immune component Ficolin 3 (Hakata Antigen) mediates the clearance of late apoptotic cells. Arthritis & Rheumatism 2007, 56: 1598
4. Thiel, S; Complement activating soluble pattern recognition molecules with collagen-like regions, mannan-binding lectin, ficolins and associated proteins. Mol Immunol 2007, 44: 3875
5. Runza, V et al; Ficolins: novel pattern recognition molecules of the innate immune response. Immunobiol 2008, 213: 297
6. Michalski, M et al; H-ficolin (ficolin-3) concentrations and FCN3 gene polymorphism in neonates. Immunobiology 2012