| Description | Purity>95% by SDS-PAGEExtinction CoeffA280 nm = 1.27 at 1.0 mg/mLGeneral DescriptionFactor Ba is the fragment of complement factor B that results from activation of the alternative pathway. Prepares the factor Ba fragment from factor B which was purified from normal human serum. Complement factorPurity>95% by SDS-PAGEExtinction CoeffA280 nm = 1.27 at 1.0 mg/mLGeneral DescriptionFactor Ba is the fragment of complement factor B that results from activation of the alternative pathway. Prepares the factor Ba fragment from factor B which was purified from normal human serum. Complement factor B is a glycosylated protein composed of a single 93,000 Da polypeptide chain. Factor B is anessential component of the alternative pathway of complement activation and is found in plasma at approximately 200 µg/mL. In the presence of Mg++ factor B binds to C3b and the C3b,B complex can be activated by factor D, a serine protease that circulates as an active trypsin-like serine protease. Cleavage of factor B by factor D causes the release of the Ba fragment (33,000 Da) and leaves the 60,000 Bb fragment bound to C3b. This Ba fragment comes from the N-terminal of factor B and it contains three CCP domains which interact with C3b (Morley, B.J. and Walport, M.J. (2000)). The isolated fragment Ba has been reported to have a weak affinity for C3b and to inhibit the interaction of factor B with C3b thus inhibiting the activation of the alternative pathway (Pryzdial, E.L.and Isenman, D.E., (1987)).The fragments of factor B (Ba and Bb) have been proposed to elicit several biological responses. See the section titled Function below and in the product description for the Bb fragment.FunctionThe fragments of factor B (Ba and Bb) have been proposed to elicit numerous biological responses; however, many of these activities have proved to be controversialwith an inconsistent record of reproducibility. It is not yet clear whether these failures are due to different experimental conditions, more highly purified Ba and Bb than available in the early days or the need to test fresh, in situ-prepared fragments, as has been suggested. Both fragments Ba and Bb have been reported to bind to B lymphocyte receptors and modulate proliferation (Kolb, W.P., et al. (1989)). Ba, but not Bb, was shown to exhibit growth-supporting activity for activated murine B lymphocytes (Praz, F. and Ruuth, E. (1986)). On the other hand, Ba has been reported to inhibit human B cell prolifieration (Ambrus, J.L. et al. (1990)). The small fragment Ba has been reported to show chemotactic activity with neutrophils and macrophages, but this effect is so much lower than that of C5a or even C5adesArg that its effect in vivo may be negligible (Morgan, B.P., (1990)). In a study of smooth muscle contractile activity, neither Ba nor Bb caused contraction of guinea pig ileum (a sensitive test for C5a) or histamine release from rat mast cells, but Ba caused guinea pig PMN to increase chemotactic activity (Hamuro, J., et al. (1978)).AssaysThere are no convenient assays for the biological activity of Ba. Several companies produce ELISA kits for measuring Ba levels in blood samples (Dodds, A.W. and Sim, R.B. (1997)).ApplicationsSplit products of factor B in plasma are indicative of activation of the alternative pathway in vivo. ELISA kits for measurement of Ba and Bb are commercially available. These have been used in numerous human and animal studies (Lynch, A.M., et al. (2008)). See In vivo section below.In vivoThe average concentration of factor B in blood is 200 µg/mL (range 170-258 µg/mL) in human plasma. Factor B is an acute phase protein whose plasma levels increases during inflammation. The fragments Ba and Bb are released upon activation of the alternative pathway and circulate in blood until cleared (Lynch, A.M., et al. (2008)). Baseline levels in normals have been reported to be 29 ng Ba/mL (0.015% of the factor B concentration). The concentration of fragment Ba in patients with tubular proteinuria was found to be elevated more than 100-fold (4800 ng/mL)(Oppermann, M. et al. (1991)). There was a minimal increase in the plasma concentration of fragment Bb in these patients.GeneticsThe gene for factor B is located on human chromosome 6p21.3 within the MHC class III region between the class I and class II regions. The factor B gene lies between the larger gene for C2 (to which it is highly homologous) and genes for C4A and C4B. The gene is composed of 18 exons and spans 6 kb.DeficienciesNo natural deficiencies of factor B have been identified in humans or animals.Mice deficient in factor B (B-/- mice), compared to wild-type, exhibit much lower or no pathology in a wide variety of diseases where alternative pathway activation is the cause of or exacerbates the pathology of these diseases. See Diseases section below. Acquired and secondary deficiencies do occur in humans. Human factor I deficiencies exhibit very low factor B levels due to the fact that C3b is not inactivated in the absence of factor I and C3b accumulates in blood. This results in binding of factor B, cleavage by factor D and rapid release of fragments Ba and Bb. Transfusions with normal plasma or reconstitution with factor I temporarily stop or slow production of these fragments.Precautions/Toxicity/HazardsThis protein is purified from human serum and therefore precautions appropriate for handling any blood-derived product must be used even though the source was shown by certified tests to be negative for HBsAg, HTLV-I/II, STS, and for antibodies to HCV, HIV-1 and HIV-II.Hazard Code: B WGK Germany 3MSDS available upon request... Read More | Protein Purity>95% by SDS-PAGEExtinction Coeff.A276 nm = 0.456 at 1.0 mg/mLMolecular Weight8,759 Da (single chain)General DescriptionNatural human C4a is prepared by cleavage of human C4 protein by human C1s. It is produced during activation of both the classical and lectin pathways of complementProtein Purity>95% by SDS-PAGEExtinction Coeff.A276 nm = 0.456 at 1.0 mg/mLMolecular Weight8,759 Da (single chain)General DescriptionNatural human C4a is prepared by cleavage of human C4 protein by human C1s. It is produced during activation of both the classical and lectin pathways of complement. C4a is a member of the anaphylatoxin family of three proteins (C3a, C4a and C5a) produced by the activation of complement (Hugli, T.E. et al. (1981)). It is an unglycosylated polypeptidecontaining 77 amino acids with a molecular mass of 8,759 daltons. Many of the biological functions of C4a are similar to those of C3a, but the specific activities are far below those of C3a. C4a activity is so low, in fact, that it was initially thought to be inactive. These measured activities include inducing muscle contraction in the guinea pig ileum test (spasmogenic activity), desensitization of muscle to C3a stimulation suggesting that the same receptor for both C3a and C4a is involved (tachyphylactic activity) and inducing vascular permeability in human skin (Gorski J.P. et al. (1979)). C4a does not show tachyphylactic activity against C5a or chemotactic activity. Removal of the C-terminal arginine by serum carboxypeptidase N destroys all these activities (Meuller-Ortiz, S.L., et al. (2009)). C4a appears to act through the C3a receptor (C3aR) which is a G-protein coupled receptor found widely distributed on peripheral tissues, lymphoid cells (neutrohphils, monocyes, and eosinophils) and in the central nervous system (astrocytes, neurons and glial cells) (Law, S.K.A. and Reid, K.B.M. (1995)). Physical Characteristics & StructureMolecular weight: 8,759 calculated molecular mass. Observed mass (MALDI-TOF) is 8,762 + 9 mass units. pI = 9.0 to 9.5 (Gorski, J.P. et al. (1981))Amino acid sequence (77 amino acids): NVNFQKAINE KLGQYASPTA KRCCQDGVTR LPMMRSCEQR AARVQQPDCR EPFLSCCQFA ESLRKKSRDK GQAGLQRC4a is thought to be structurally very similar to C3a and C5a to which it is homologous. Thus its 3D structure is probably similar to the X-ray-derived crystal structureof C3a (Huber, R. et al. (1980)) and the NMR derived structure of C3a: Nettesheim, D.G. et al. (1988); Murray, I. et al. (1999).FunctionSee General Description above. C4a exhibits much weaker biological activities than C3a and C5a. Its activity in inducing erythema and edema in human skin is 25,000-fold weaker than that of C5a and 100-fold weaker than C3a per nanomole. The spasmogenic activity of C4a is 2000-fold weaker than C5a and 100-fold weaker than that of C3a. Due to these differences the role of C4a in these responses in vivo is thought to be negligible.AssaysTwo well established assays for C4a and C3a functional activities include induction of contraction in the guinea pig ileum and the permeation of a dye such as trypan blue from the vasculature into skin. The anaphylatoxins also induce mast cell degranulation, (measured as histamine release), platelet aggregation, IL-1 release from monocytes and the release of prostaglandins and leukotrienes from many cells and tissues. The other assays used for C3a (Dodds, A.W. and Sim, R.B. (1997)) should also respond to C4a, but few reports have described utilizing these assays with C4a. ELISA kits for the assay of C4a levels (or more correctly C4a desArg levels) in blood and other fluids are sold by several companies. These measurements are useful for detecting complement activation in vivo, but the interpretation of their meaning is complicated by the fact that clearance of the anaphylatoxins is rapid. In vivoFreshly drawn normal human serum contains significant levels of all three anaphylatoxins. Although these may represent the resting concentration in vivo it is difficult to draw or store blood without some complement activation so a true in vivo concentration is difficult to determine. The presence of EDTA and Futhan in the collection tubes can minimize this background (Pfeifer, P.H. et al. (1999)). Full activation of all C4 in blood (600µg/mL) would result in ~3,400 nM C4a (~30 µg/mL). Due to the low biological activity of C4a it could require activation of most of the C4 in a small region to achieve the micromolar C4a concentrations necessary to elicit a response.RegulationC4a levels are regulated by three processes: formation, inactivation and clearance. There are two enzymes that cleave C4 and release C4a: C1s and MASP-2. C4a is “inactivated” by removal of its C-terminal arginine amino acid. The product C4a desArg (or C4a without the C-terminal arginine) is produced by the action of the plasma enzyme carboxypeptidase N (Mueller-Ortiz S.L. et al. (2009)). The inactivation is rapid and most C4a is converted to C4a desArg within minutes of its formation. Inactivated C4a lack measurable biological activity. Because of the large number of cells bearing C3a/C4areceptors (endothelial, immune, smooth muscle, neuronal, etc.) the capture, internalization and digestion of C4a and C4a desArg probably results in its removal from circulation.DeficienciesA deficiency of C4 or a deficiency of all of the enzymes that cleave C4 to generate C4a could result in the absence of C4a. There are no known complete deficiencies of all ofthe C4 cleaving enzymes. Examples of C4 deficient humans and mice exist (Wessels, M.R. et al. (1995)), but the degree to which pathologies associated with C4 deficiency are due to the lack of C4 or the absence of C4a is unclear. DiseasesThere are no known diseases connected to C4a or C4a desArg. Precautions/Toxicity/HazardsThe source of C4a is human serum, therefore appropriate precautions must be observed even though the source was shown by certified tests to be negative for HBsAg, HTLV-I/II, STS, and for antibodies to HCV, HIV-1 and HIV-II.Injection can cause anaphylatic shock which is a generalized circulatory collapse similar to that caused by an allergic reaction.Hazard Code: B WGK Germany 3... Read More | Inquire | Source: Microorganism Isoelectric point: 6.5 Michaelis constant: 9.2×10^-3 M (D-Glucose); 8.6×10^-3 M (NAD) Optimum pH: 9.0~9.5 Fig. 1Optimum temperature: 55℃ Fig. 3pH Stability: 6.0-10.0 (25℃, 24hr) Fig. 2Thermal stability: <50℃ (pH 8.0, Source: Microorganism Isoelectric point: 6.5 Michaelis constant: 9.2×10^-3 M (D-Glucose); 8.6×10^-3 M (NAD) Optimum pH: 9.0~9.5 Fig. 1Optimum temperature: 55℃ Fig. 3pH Stability: 6.0-10.0 (25℃, 24hr) Fig. 2Thermal stability: <50℃ (pH 8.0, 30min) Fig. 4Inhibitors: NEM,SDS Effect of various chemicals: Table 1Reaction:... Read More | Biochemical Test:Electrophoresis (purity > 80%) |