Cat. # | Size | Price | Inventory |
---|---|---|---|
34920S | 100 µl (50 tests) |
REACTIVITY | M |
SENSITIVITY | Endogenous |
MW (kDa) | |
Source/Isotype | Rabbit IgG |
Product Information
Application | Dilution |
---|---|
Immunofluorescence (Frozen) | 1:50 - 1:200 |
Immunofluorescence (Immunocytochemistry) | 1:400 - 1:3200 |
Flow Cytometry (Fixed/Permeabilized) | 1:50 |
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Cover sections with 4% formaldehyde dilute in 1X PBS.
NOTE: Formaldehyde is toxic, use only in 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: 222
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
小鼠
大鼠, 仓鼠
使用与小鼠 PD-1 蛋白的 Ala242 周围的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。
程序性细胞死亡 1 蛋白 (PD-1, PDCD1, CD279) 是免疫受体 CD28 家族的一员,该家族可调控 T 细胞激活和免疫应答 (1-3)。PD-1 蛋白有一个胞外 Ig V 结构域、一个跨膜结构域和一个包含免疫受体酪氨酸抑制基序 (ITIM) 和一个免疫受体酪氨酸转换基序 (ITSM) 的细胞浆尾区。细胞表面配体 PD-L1 和 PD-L2 可激活 PD-1 (4)。激活时,PD-1 ITIM 和 ITSM 磷酸化会导致蛋白酪氨酸磷酸酶 SHP-1 和 SHP-2 的募集,从而抑制 TCR 信号转导 (5-7)。除了激活的 T 细胞,PD-1 还在激活的 B 细胞和单核细胞中表达,尽管它在这些细胞类型中的作用并不完全确定 (8)。PD-1 通路在免疫耐受性方面发挥重要作用 (3),但研究表明癌细胞通常利用这个通路来逃避免疫监视 (9)。因此,PD-1 及其配体的阻断被证明是一个极好的肿瘤预防策略 (10)。
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