Description
The RmC7H8 monoclonal antibody specifically recognizes the C1q component of the mouse macromolecular C1 complex. The complex is comprised of C1q and 2 molecules each of the serine proteases, C1r and C1q. The C1 macromolecular complex, C1qC1r2C1s2, is bound together by Ca2+ ions. C1q is a serum protein that is synthesized by macrophages and microglia. It exists as an ~460 kDa protein formed from 18 polypeptide chains comprised of three different subunits named C1qa, C1qb, and C1qc. Each chain contains an N-terminal collagen-like sequence and a C-terminal globular gC1q module. Structural studies reveal that C1q is formed as a hexamer with 6 collagen-like triple helices, forming a central fiber bundle, each extended by globular domains. C1q is a multifunctional protein that can regulate a variety of cellular processes in addition to activating the classical complement pathway (CCP). C1q globular regions mediate target recognition such as binding to the Fc regions of IgG and IgM antibodies found in immune complexes including antibodies bound to pathogens or target cells and subsequent activation of the CCP. The C1q globular regions can also bind to bacterial and viral surface proteins as well as altered self elements including histones, DNA, and annexins on the surface of apoptotic or necrotic cells. The enhanced phagocytosis mediated through interaction of bound C1q with various receptors expressed by phagocytes, possibly combined with further complement activation and opsonization of cells, may contribute to the safe removal of stressed or dead cells that are pro-inflammatory. Conformational changes in target bound C1q's collagen-like regions can lead to interaction with and activation of the C1r and C1s proteases which result in activation of the CCP.
Bead-based compensation or unmixing controls, such as BD® CompBeads or BD™ SpectraComp™, can be used as surrogates to assess fluorescence spillover when bound to fluorochrome-conjugated antibodies. Although these beads have spectral properties similar to cells, variations in spectral emission may occur, resulting in differing spillover values compared to biological controls. Therefore, it is considered best practice to compare the spillover obtained from cells and bead-based controls when using BD® CompBeads or BD™ SpectraComp™ for the first time, to ensure they are appropriate for the intended application.
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to the dye under optimum conditions that minimize unconjugated dye and antibody.
1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
2. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
3. For U.S. patents that may apply, see bd.com/patents.
4. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
5. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
6. The production process underwent stringent testing and validation to assure that it generates a high-quality conjugate with consistent performance and specific binding activity. However, verification testing has not been performed on all conjugate lots.
7. When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
8. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
9. An isotype control should be used at the same concentration as the antibody of interest.
10. Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.
11. Please observe the following precautions: We recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to protect exposure of conjugated reagents, including cells stained with those reagents, to any room illumination. Absorption of visible light can significantly affect the emission spectra and quantum yield of tandem fluorochrome conjugates.
12. Tandem fluorochromes contain both an energy donor and an energy acceptor. Although every effort is made to minimize the lot-to-lot variation in the efficiency of the fluorochrome energy transfer, differences in the residual emission from the donor may be observed. Additionally, multi-laser cytometers may directly excite both the donor and acceptor fluorochromes. Therefore, we recommend for every tandem conjugate, a matched individual single-stain control be acquired for generating a compensation or spectral unmixing matrix.
