Preparation and Storage
推奨アッセイ手順
Because CD179b is expressed at very low levels on bone-marrow-derived early B lineage cells, we recommend the use of Mouse BD Fc Block™ purified anti-mouse CD16/CD32 mAb 2.4G2 (Cat. no. 553141/553142) and amplification of staining by the use of a "bright" second-step reagent, such as Streptavidin-PE (Cat. no. 554061) or Streptavidin-APC (Cat. no. 554067). It is difficult to distinguish the CD179b+ cells among the total bone-marrow population. Therefore, the staining protocol described by Winkler, et al., should be used. Essential features of this procedure include immunomagnetic depletion of surface IgM+ cells and incubation of the cells at 37°C for 1 hour in tissue culture medium.
Product Notices
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- An isotype control should be used at the same concentration as the antibody of interest.
- 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.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
関連製品
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The pre-B cell receptor (pre-BCR) expressed during the early stages of B lymphocyte development is a heterodimer of immunoglobulin heavy chain (IgH) with surrogate light chain, which is an Ig-light-chain-like molecule composed of the non-covalently linked CD179b (λ5) and CD179a (VpreB) proteins. The pre-BCR is believed to control IgH repertoire selection and proliferation of differentiating B lymphocytes. The LM34 antibody reacts with λ5 and surrogate light chain, but not VpreB alone, in transfected X63-Ag8.653 cells. It detects surrogate light chain on pro-B and pre-B cell lines, in the presence or absence of IgH, but not on IgM-positive B lymphocytes. It also detects surrogate light chain or λ5 on the surface and in the cytoplasm of pro-B or pre-B cells from the bone marrow of normal or RAG2-deficient mice. It has been noted that the (CD45R/B220) cell-surface expression of surrogate light chain is upregulated after a one-hour incubation of bone-marrow leukocytes in tissue culture medium at 37°C. At the earliest stages of B lymphopoiesis, before IgH is available, the surrogate light chain associates with a complex of glycoproteins, including a nonclassical cadherin, which could be involved in selective adhesion events during B-lymphocyte development. After immunization, cell-surface λ5 is detected on a subset of splenic Ig light chain-positive germinal-center B lymphocytes.
Development References (9)
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Karasuyama H, Rolink A, Melchers F. A complex of glycoproteins is associated with VpreB/lambda 5 surrogate light chain on the surface of mu heavy chain-negative early precursor B cell lines. J Exp Med. 1993; 178(2):469-478. (Immunogen). View Reference
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Karasuyama H, Rolink A, Shinkai Y, Young F, Alt FW, Melchers F. The expression of Vpre-B/lambda 5 surrogate light chain in early bone marrow precursor B cells of normal and B cell-deficient mutant mice. Cell. 1994 April; 77(1):133-143. (Biology). View Reference
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Martensson IL, Ceredig R. Review article: role of the surrogate light chain and the pre-B-cell receptor in mouse B-cell development. Immunology. 2000; 101(4):435-441. (Biology). View Reference
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Meffre E, Papavasiliou F, Cohen P, et al. Antigen receptor engagement turns off the V(D)J recombination machinery in human tonsil B cells. J Exp Med. 1998; 188(4):765-772. (Biology). View Reference
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Melchers F, ten Boekel E, Seidl T, et al. Repertoire selection by pre-B-cell receptors and B-cell receptors, and genetic control of B-cell development from immature to mature B cells. Immunol Rev. 2000; 175:33-46. (Biology). View Reference
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Ohnishi K, Shimizu T, Karasuyama H, Melchers F. The identification of a nonclassical cadherin expressed during B cell development and its interaction with surrogate light chain. J Biol Chem. 2000; 275(40):31134-31144. (Biology). View Reference
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Rolink A, Grawunder U, Winkler TH, Karasuyama H, Melchers F. IL-2 receptor alpha chain (CD25, TAC) expression defines a crucial stage in pre-B cell development. Int Immunol. 1994; 6(8):1257-1264. (Biology). View Reference
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Shimizu T, Mundt C, Licence S, Melchers F, Martensson IL. VpreB1/VpreB2/lambda 5 triple-deficient mice show impaired B cell development but functional allelic exclusion of the IgH locus. J Immunol. 2002; 168(12):6286-6293. (Biology). View Reference
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Winkler TH, Rolink A, Melchers F, Karasuyama H. Precursor B cells of mouse bone marrow express two different complexes with the surrogate light chain on the surface. Eur J Immunol. 1995; 25(2):446-450. (Biology). View Reference
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