Cat. # | Size | Price | Inventory |
---|---|---|---|
57761S | 100 µl (50 tests) |
REACTIVITY | H |
SENSITIVITY | Endogenous |
MW (kDa) | |
Source/Isotype | Rabbit IgG |
Product Information
Application | Dilution |
---|---|
Immunofluorescence (Immunocytochemistry) | 1:50 - 1:200 |
Flow Cytometry (Fixed/Permeabilized) | 1:50 |
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
NOTE: Cells should be grown, treated, fixed and stained directly in multi-well plates, chamber slides or on coverslips.
Aspirate liquid, then cover cells to a depth of 2–3 mm with 4% formaldehyde diluted in 1X PBS.
NOTE: Formaldehyde is toxic, use only in a fume hood.
NOTE: All subsequent incubations should be carried out at room temperature unless otherwise noted in a humid light-tight box or covered dish/plate to prevent drying and fluorochrome fading.
posted November 2006
revised November 2013
Protocol Id: 182
All reagents required for this protocol may be efficiently purchased together in our Intracellular Flow Cytometry Kit (Methanol) #13593, or individually using the catalog numbers listed below.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
NOTE: When including fluorescent cellular dyes in your experiment (including viability dyes, DNA dyes, etc.), please refer to the dye product page for the recommended protocol. Visit www.cellsignal.com for a full listing of cellular dyes validated for use in flow cytometry.
NOTE: Adherent cells or tissue should be dissociated and in single-cell suspension prior to fixation.
NOTE: Optimal centrifugation conditions will vary depending upon cell type and reagent volume. Generally, 150-300g for 1-5 minutes will be sufficient to pellet the cells.
NOTE: If using whole blood, lyse red blood cells and wash by centrifugation prior to fixation.
NOTE: Antibodies targeting CD markers or other extracellular proteins may be added prior to fixation if the epitope is disrupted by formaldehyde and/or methanol. The antibodies will remain bound to the target of interest during the fixation and permeabilization process. However, note that some fluorophores (including PE and APC) are damaged by methanol and thus should not be added prior to permeabilization. Conduct a small-scale experiment if you are unsure.
NOTE: Count cells using a hemocytometer or alternative method.
posted July 2009
revised June 2020
实验步骤编号:407
人
使用人雌激素受体 α 蛋白氨基末端特异性重组蛋白对动物进行免疫接种来产生单克隆抗体。
雌激素受体 α (ERα) 是类固醇受体超家族的一员,它包含高度保守的 DNA 结合域和配体结合域 (1)。通过其雌激素非依赖性和雌激素依赖性激活域(分别为 AF-1 和 AF-2),ERα 通过召集共激活因子蛋白并与一般转录机制相互作用来调节转录 (2)。多个位点的磷酸化是调节 ERα 活性的一个重要机制 (3-5)。Ser104、106、118 和 167 位于氨基末端转录激活功能域 AF-1,这些丝氨酸残基的磷酸化在调节 ERα 活性方面起着重要作用。Ser118 可能是转录调节性激酶 CDK7 的底物 (5)。Ser167 可能被 p90RSK 和 Akt 磷酸化 (4,6)。研究文献研究显示,Ser167 的磷酸化可能会使乳腺癌患者对他莫昔芬产生耐药性 (4)。
探索与本品相关的通路。
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