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Catalog # HK359

MAp44, Human, ELISA kit

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The complement system is an essential part of the innate immune response. It consists of more than 30 proteins which act as a proteolytic cascade as first defense against pathogens. Complement modulates also B- and T-cell responses thereby bridging innate and adaptive immunity. Complement is activated in three ways, the classical, alternative and lectin pathways. The lectin pathway is initiated by pathogen-recognition receptors mannan-binding lectin (MBL), H-ficolin, L-ficolin, M-ficolin or collectin-K1 which recognize a broad range of pathogens or apoptotic cells. All molecules form complexes with MBL-associated serine proteases (MASPs). Five MASP proteins have thus far been identified: MASP1, MASP2, MASP3 and their truncated proteins MAp44 (also called MAp1) and MAp19 (or sMAP). MASP1&3 and MAp44 are all derived from MASP1 gene. MAp44 mRNA consists of the first 8 exons encoding the first four domains identical to those of MASP-1 and MASP-3 and an extra exon encoding 17 amino acids unique to MAp44. Deficiencies in the lectin pathway are associated with sensitivity for infection and the lectin-MASP complex is also involved in activation of the coagulation system. The function of MASP3, MAp44 and MAp19 in the lectin pathway are largely unknown. Competition of recombinant MAp44 in complex with either recombinant MBL or recombinant H-ficolin causes decreased levels of cleavage of C4 in sera depleted of MBL and H-ficolin, respectively, suggesting that MAp44 has an inhibition effect on activation of complement system. MAp44 may represent a tissue-specific protective mechanism by which local inflammation is prevented. MBL-associated protein MAp44 has shown cardio-protective effects in murine models. Although associated with cardiovascular risk factors, MAp44 levels were not associated with severity of disease in humans. In human plasma concentrations are 0.8-3.2 µg/ml. The assay captures MAp44 with a monoclonal antibody recognizing the C-terminus and uses a detection antibody reacting with the CCP-1 domain.

Application
The human MAp44 ELISA kit is to be used for the in vitro quantitative determination of human MAp44 in serum and plasma samples.

Principle
The human MAp44 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 a plate with twelve disposable 8-well strips allows free choice of batch size for the assay. Samples and standards are incubated in microtiter wells coated with antibodies recognizing human MAp44. Biotinylated tracer antibody will bind to the captured human MAp44. 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 human MAp44standards (log). The human MAp44 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 citrate plasma sample containing human MAp44. The diluted sample was measured in the assay.

Recovery
Normal human blood samples (plasma) containing baseline levels of human MAp444, were spiked with human MAp44, in concentrations of 50 and 5 ng/ml. Samples with and without human MAp44, were incubated for 1 hour at room temperature. Samples were measured using the ELISA. Values for human MAp44, ranged between 83% and 99%.

Product type
Assays
Quantity
1 x 96 det., 2 x 96 det.
Standard range
1.6 to 100 ng/ml
Detection level
1.6 ng/ml
Working volume
100 µl/well
Species
Human
Cross reactivity
Horse – Yes, Mouse – No, Pig – No, Rabbit – No, Rat – No
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
Cardiology and metabolism, Infectious diseases
Safety Data Sheet Assay Manual
HK359
368 kb

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