Preparation and Storage
推奨アッセイ手順
Applications include immunoprecipitation, western blot analysis (1-2 µg/ml) and electrophoretic mobility band shift assays (EMSA). Positive controls: E2A proteins (E12 and E47) have been detected in cells derived from both B-cell and T-cell lineages including the pre B-cell line Nalm-6, the mature B-cell line Namalwa (ATCC CRL-1432), the T-cell line Jurkat (ATCC TIB-152). E2A has also been detected in HeLa cervical carcinoma cells (ATCC CCL-2).
Product Notices
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- 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.
関連製品
B lymphoid cells go through many intermediate steps before becoming plasma ells whichproduce and secrete immunoglobulins (Ig). The earliest stage of B-cell differentiate is represented by pro-B lymphocytes that contain both the Ig heavy- and light-chain genes in the transcriptionally inactive germ line cnfiguration. As ro-B lymphocytes differentiate into pro-B cells, Ig heavy-chain genes rearrange and get transcribed and translated. However, the Ig lightchain genes are not transcribed before the pre-B cells differentiate into mature B-lymphocytes. The developmental regulation of the Ig gene expression is dependent on various sequences in the Ig enhancer region. One class of such regulatory sequence elements comprises the so-called E-boxes which share the NNCANNTGNN consensus sequence. The E2 boxes are particularly interesting because they are present in muscle and pancreas-specific enhancers. A family of proteins binds to the E2 box. These proteins share a common amino acid sequence motif that is proposed to form two amphipathic helices interrupted by a loop, designated the helix-loop-helix (HLH) motif. The HLH motif mediates homo- as well as heterodimerization with other HLH proteins. Most HLH proteins possess a basic region located N terminal of the HLH region which is responsible for DNA binding. Two E2 box binding proteins have been described (E12 and E47) that arise as alternatively spliced form of the E2A gene. E12 and E47 are identical except in the HLH region. These two proteins have been directly implicated in the regulation of B-cell, muscle and pancreas-specific gene expression. The E2-2 gene product is closely related to the E2A gene products but is encoded by a separate gene. E47 migrates at a reduced molecular weight of ~75 kDa. G98-271 reacts specifically with human E12 and E47 protein, alternatively spliced products of the E2A gene. It does not cross-react with the E2-2 protein. The specificity of the antibody was tested by immunoprecipitation and electrophoretic mobility band shift assays (EMSA) using recombinant protein (E2-2, E12, and E4 7). The HLH motif as well as flanking regions of recombinant human E12 potein was used as the immunogen.
Development References (7)
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Bain G, Gruenwald S, Murre C. E2A and E2-2 are subunits of B-cell-specific E2-box DNA-binding proteins.. Mol Cell Biol. 1993; 13(6):3522-3529. (Clone-specific: Immunoprecipitation, Western blot). View Reference
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Bain G, Robanus Maandag EC, te Riele HP. Both E12 and E47 allow commitment to the B cell lineage. Mol Cell Biol. 1993; 6(2):145-154. (Clone-specific: Western blot). View Reference
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Buskin JN, Hauschka SD. Identification of a myocyte nuclear factor that binds to the muscle-specific enhancer of the mouse muscle creatine kinase gene. Mol Cell Biol. 1989; 9(6):2627-2640. (Biology). View Reference
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Lenardo M, Pierce JW, Baltimore D. Protein-binding sites in Ig gene enhancers determine transcriptional activity and inducibility. Science. 1987; 236(4808):1573-1577. (Biology). View Reference
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Murre C, McCaw PS, Baltimore D. A new DNA binding and dimerization motif in immunoglobulin enhancer binding, daughterless, MyoD, and myc proteins. Cell. 1989; 56(5):777-783. (Biology). View Reference
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Sun XH, Baltimore D. An inhibitory domain of E12 transcription factor prevents DNA binding in E12 homodimers but not in E12 heterodimers. Cell. 1991; 64(2):459-470. (Biology). View Reference
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Whelan J, Cordle SR, Henderson E, Weil PA, Stein R. Identification of a pancreatic beta-cell insulin gene transcription factor that binds to and appears to activate cell-type-specific expression: its possible relationship to other cellular factors that bind to a common insulin gene sequence. Mol Cell Biol. 1990; 10(40):1564-1572. (Biology). View Reference
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