Cat. # | Size | Price | Inventory |
---|---|---|---|
72800S | 100 µl (50 tests) |
REACTIVITY | H |
SENSITIVITY | Endogenous |
MW (kDa) | |
Source/Isotype | Rabbit IgG |
Product Information
Application | Dilution |
---|---|
Immunofluorescence (Immunocytochemistry) | 1:800 |
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
使用与人 p14 ARF 蛋白中 Pro126 周围残基相对应的合成肽,对动物进行免疫接种来产生单克隆抗体。
人 p14 ARF (小鼠 p19 ARF)是一种在人肿瘤细胞中频繁失活的促凋亡细胞周期调节分子 (1)。在大多数细胞类型中,p14 ARF 的基础表达较低,但该蛋白在癌基因表达的刺激下会出现聚集 (2,3)。p14 ARF 水平升高可促进 MDM2 降解,从而导致 p53 蛋白水平升高及后续的细胞周期停滞和/或凋亡 (4)。虽然多数 p14 ARF 信号转导在传统意义上被认为是具有 p53 依赖性,但近来更多报道描述了 p53 非依赖性 p14 ARF 信号转导会导致细胞周期停滞和凋亡 (5,6)。
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