Cat. # | Size | Price | Inventory |
---|---|---|---|
26632T | 20 µl | ||
26632S | 100 µl |
REACTIVITY | H M R |
SENSITIVITY | Endogenous |
MW (kDa) | 14, 16 |
Source/Isotype | Rabbit IgG |
Product Information
Application | Dilution |
---|---|
Western Blotting | 1:1000 |
Immunofluorescence (Immunocytochemistry) | 1:200 |
Flow Cytometry (Fixed/Permeabilized) | 1:100 |
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Loading of prestained molecular weight markers (#59329, 10 µl/lane) to verify electrotransfer and biotinylated protein ladder (#7727, 10 µl/lane) to determine molecular weights are recommended.
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
Recommended Fluorochrome-conjugated Anti-Rabbit secondary antibodies:
NOTE: Cells should be grown, treated, fixed and stained directly in multi-well plates, chamber slides or on coverslips.
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 December 2010
Protocol Id: 3
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:404
人, 小鼠, 大鼠
通过采用与人 GABARAPL1 蛋白的氨基末端附近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。
GABAA 受体相关蛋白 (GABARAP) 是在自噬中具有关键作用的 Atg8 家族蛋白,该蛋白质最初作为与调节受体向质膜转运的 GABAA 受体结合的蛋白质发现 (1)。该家族中的蛋白质(包括微管相关蛋白轻链 3 (LC3) 和 GATE-16 (GABARAPL2))在自噬性刺激下(如饥饿后)并入自噬体膜 (2)。如同其他家族成员,GABARAP 在其羧基末端受剪切,并进而导致借助磷脂类磷脂酰乙醇胺或磷脂酰丝氨酸接合 (3,4)。这种加工过程将 GABARAP 从 I 型转化成涉及自噬体生物生成的 II 型膜结合形式。GABARAP 加工包括 Atg4 家族成员剪切 (5,6) ,随后是 E1 和 E2 接合(例如酶 Atg7 和 Atg3 )(7,8)。鉴定 GABARAPL1/GEC1(与 GABARAP 高度相关的蛋白质)为雌激素诱导型基因,并且它还与自噬体结合 (9-11)。
与其他家族成员 (12-14) 相比,在 CNS 中 γ-氨基丁酸受体相关蛋白样 1 (GABARAPL1) 表达程度似乎更高。GABARAPL1 的表达与一些癌症(包括肝细胞癌和乳腺癌)的预后相关 (15,16)。抑制乳腺癌细胞中 GABARAPL1 表达可减弱自噬通量,导致代谢变化并产生促癌活性 (17)。
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