| Description | APOH Human Pre-designed siRNA Set A contains three designed siRNAs for APOH gene (Human), as well as a negative control, a positive control, and a FAM-labeled negative control. Components APOH siRNA-1: 5 nmol (HPLC) APOH siRNA-2: 5 nmol (HPLC) APOH siRNA-3: 5 nmol (HPLC) siRNA Negative Control: 5 APOH Human Pre-designed siRNA Set A contains three designed siRNAs for APOH gene (Human), as well as a negative control, a positive control, and a FAM-labeled negative control. Components APOH siRNA-1: 5 nmol (HPLC) APOH siRNA-2: 5 nmol (HPLC) APOH siRNA-3: 5 nmol (HPLC) siRNA Negative Control: 5 nmol (HPLC) FAM-labeled siRNA Negative Control: 5 nmol (HPLC) GAPDH siRNA Positive Control:5 nmol (HPLC)... Read More | C1q separated from C1r and C1s and from other stabilizing proteins tends to aggregate easily. Because it was isolated and studied in numerous research laboratories, many buffers have been used to stabilize concentrated C1q and prevent aggregation. About half of the scientists prefer high salt and C1q separated from C1r and C1s and from other stabilizing proteins tends to aggregate easily. Because it was isolated and studied in numerous research laboratories, many buffers have been used to stabilize concentrated C1q and prevent aggregation. About half of the scientists prefer high salt and the other prefer 40% glycerol in the storage buffer.C1q is purified from pooled normal human plasma. C1q is part of the C1 complex and this complex is the first complement component in the cascade referred to as the classical pathway of complement. C1 is actually a non-covalent assembly of three different proteins (C1q, C1r, and C1s) bound together in a calcium-dependent complex. C1q has six extended arms with domains at the end of each arm that bind to the Fc domains of immunoglobulins. When antibodies bind to antigens forming immune complexes they cluster allowing two or more of its six arms of C1q to bind to the Fc domains of antibodies such as IgG or IgM. The binding of multiple arms to immune complexes causes the two C1r proteins in the complex (protease zymogens) to auto-activate producing two C1r proteases that cleave and activate the two C1s protease zymogens in the complex. Activated C1s cleaves complement component C4 releasing C4a and initiating covalent attachment of C4b to the activating surface. Activated C1s also cleaves C2 and the larger fragment of C2 binds to the surface-attached C4b forming C4b,C2a which is the C3/C5 convertase of the classical pathway.Extinction Coeff.A₂₈₀ nm = 0.68 at 1.0 mg/ml for pure C1q Molecular weight:410,000 Da (18 chains)Preservative:None, 0.22 µm filtered.Source:Normal human serum (shown by certified tests to be negative for HBsAg, HTLV-I/II, STS, and for antibodies to HCV, HIV-1 and HIV-II).Physical Characteristics & StructureC1q is a high molecular weight complex of 18 polypeptide chains. Each of the six arms of C1q contains three chains, an A chain (26,000 daltons), a B chain (25,000 daltons) and a C chain (24,000 daltons). The three chains are coiled into a collagen-like triple helix over approximately half their length. Half of this collagen region forms a central core where all 18 chains come together. The chains are joined in this core by disulfides in the pattern A-B and C-C. There is a bend in the center of the collagen region allowing the arms to extend away from each other. Globular heads at the far ends of the collagen arms possess binding sites for Fc domains of immunoglobulins. C1 complex is composed of one C1q molecule (410,000 daltons), two C1r molecules (92,000 daltons) and two C1s molecules (86,000 daltons). The complex is stable in the presence of calcium, but easily dissociates if calcium is removed. When C1 is activated the C1r and C1s subunits are each cleaved into two chain molecules due to proteolytic activation. Thus, the SDS gel pattern of C1 is very complex. Function The biological functions of C1q are described above in the General Description and Physical Characteristics sections. C1q functional activity may be assayed using C1q-depleted serum and EA cells. These assays are extremely sensitive to C1q typically yielding 50% lysis with less than 2 ng C1q in assays measuring the lysis of EA cells. AssaysThe unit of classical pathway activity is the CH50. A similar unit, the C1qH50, is used to quantitate the activity of C1q. A C1qH50 unit is the amount of functional C1q needed to lyse 50% of 3×10^7 EA cells (antibody-sensitized sheep erythrocytes) when that amount of C1q is incubated with 5-20 µL of C1q-Dpl in GVB++ in a total volume of 500 µL for 30 min at 37℃. This amount of C1q indicates the sensitivity of the assay for C1q which is typically about 1 ng C1q with 10 µL C1q-Dpl. See the Certificate of Analysis for lot specific values.ApplicationsC1q is used to coat ELISA plates to capture and quantitate immune complexes in clinical samples. A number of commercial companies sell diagnostic kits for immune complex detection and quantitation. These kits are based on the ability of C1q to bind well to immune complexes, but to not bind significantly to monomeric immunoglobulins. GeneticsThe EMBL/Genbank cDNA accession numbers are: C1q A chain (P02745), C1q B chain (P02746), and C1q C chain (P02747). The genes for C1q chains A, B and C are all located on chromosome 1p in the order A-C-B. DeficienciesDeficiencies of each of the three components of C1 have been found. Patients lacking C1q generally have immune-complex-mediated renal disease and skin lesions. Like all patients lacking early classical pathway components C1q deficient individuals are prone to systemic lupus erythrematosis (SLE) and recurrent pyogenic infections. They lack classical pathway function and may or may not exhibit C1q antigen in blood.DiseasesSee section titled Deficiencies above. 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... 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 | Product Characteristics UNI-StabilPLUS is a universal stabilizer for the dilution and stabilization of both Horseradish Peroxidase (HRP) and Alkaline Phosphatase (AP) labeled proteins and antibodies, in order to maintain the molecular conformation and prevent loss of activity over time. This enablesProduct Characteristics UNI-StabilPLUS is a universal stabilizer for the dilution and stabilization of both Horseradish Peroxidase (HRP) and Alkaline Phosphatase (AP) labeled proteins and antibodies, in order to maintain the molecular conformation and prevent loss of activity over time. This enables the making of pre-diluted, ready-to-use conjugates, minimizing assay errors in dilution. Superior stabilization of HRP and AP conjugated antibodies in low as well as high protein dilutions is seen, when using UNI-StabilPLUS. When tested with AP conjugated antibody stability is seen as follows: • at least 3 years at 2-8 °C • at least 2 years at room temperature • at least 4 weeks at 37 °C When tested with HRP conjugated antibody stability is seen as follows: • at least 2 years at 2-8 °C • at least 1 years at room temperature • at least 2 weeks at 37 °CUNI-StabilPLUS is recommended for the dilution of antibodies directed against rabbit immunoglobulins unlike HRP-StabilPLUS (cat. no. H494387) and Antibody Enhancer (cat. no. A494276).Composition & Properties UNI-StabilPLUS is a ready-to use buffer that appears as an opaque solution. The product is based on a mild acid Tris buffer containing proprietary stabilizing components. UNI-StabilPLUS contains neither BSA, nor other material from bovine serum, no azide, mercury or other toxic components.Working Procedure 1.Make a series of dilutions of the HRP- or AP conjugated protein in UNI-StabilPLUS in order to determine the optimal dilution. 2.Run the assay as usual or store the diluted conjugated protein preferably at 2-8 °C.Tips & Tricks • Avoid using phosphate buffers for AP-conjugated antibody assays. We recommend the use of Tris/HCl, Tween as the washing buffer, instead of a PBS buffer which will reduce signal significantly. • For extended stability of HRP conjugated antibodies, HRP-StabilPLUS (cat. no. H494387) is recommended. Handling & Storage • Store solution at 2-8 °C... Read More |