重组Anti-LC3B抗体[EPR18709] - Autophagosome Marker (ab192890)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR18709] to LC3B - Autophagosome Marker
- Suitable for: IHC-P, WB, ICC/IF
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
概述
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产品名称
Anti-LC3B抗体[EPR18709] - Autophagosome Marker
参阅全部 LC3B 一抗 -
描述
兔单克隆抗体[EPR18709] to LC3B - Autophagosome Marker -
宿主
Rabbit -
经测试应用
适用于: IHC-P, WB, ICC/IFmore details
不适用于: IP -
种属反应性
与反应: Mouse, Rat, Human -
免疫原
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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阳性对照
- WB: BMDM, U-87 MG, C6 and RAW 264.7 whole cell lysates; Human brain, mouse heart, rat heart, mouse brain and rat brain lysates. ICC/IF: HeLa cells (+/- chloroquine), HAP1 cells (+/- chloroquine) (HAP1-MAP1LC3B knockout cells used as negative cell line). IHC-P: Human Cerebral Cortex tissue sections
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常规说明
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
性能
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形式
Liquid -
存放说明
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
存储溶液
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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纯度
Protein A purified -
克隆
单克隆 -
克隆编号
EPR18709 -
同种型
IgG -
研究领域
相关产品
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Recombinant Protein
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Related Products
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab192890于以下的经测试应用
“应用说明”部分 下显示的仅为推荐的起始稀释度;实际最佳的稀释度/浓度应由使用者检定。
应用 | Ab评论 | 说明 |
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IHC-P |
Use a concentration of 0.1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
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WB | (8) |
1/2000. Detects a band of approximately 14, 16 kDa (predicted molecular weight: 15 kDa).
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ICC/IF | (6) |
Use a concentration of 1 µg/ml.
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说明 |
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IHC-P
Use a concentration of 0.1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
WB
1/2000. Detects a band of approximately 14, 16 kDa (predicted molecular weight: 15 kDa). |
ICC/IF
Use a concentration of 1 µg/ml. |
靶标
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功能
Probably involved in formation of autophagosomal vacuoles (autophagosomes). -
组织特异性
Most abundant in heart, brain, skeletal muscle and testis. Little expression observed in liver. -
序列相似性
Belongs to the MAP1 LC3 family. -
翻译后修饰
The precursor molecule is cleaved by APG4B/ATG4B to form LC3-I. This is activated by APG7L/ATG7, transferred to ATG3 and conjugated to phospholipid to form LC3-II. -
细胞定位
Cytoplasm > cytoskeleton. Endomembrane system. Cytoplasmic vesicle > autophagosome membrane. LC3-II binds to the autophagic membranes. - Information by UniProt
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数据库链接
- Entrez Gene: 81631 Human
- Entrez Gene: 67443 Mouse
- Entrez Gene: 64862 Rat
- Omim: 609604 Human
- SwissProt: Q9GZQ8 Human
- SwissProt: Q9CQV6 Mouse
- SwissProt: Q62625 Rat
- Unigene: 356061 Human
see all -
别名
- ATG8F antibody
- Autophagy-related protein LC3 B antibody
- Autophagy-related ubiquitin-like modifier LC3 B antibody
see all
图片
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All lanes : Anti-LC3B antibody [EPR18709] - Autophagosome Marker (ab192890) at 1/2000 dilution
Lane 1 : Wild-type HepG2 untreated control cell lysate
Lane 2 : Wild-type HepG2 Treated Chloroquine (50 uM, 16 h) cell lysate
Lane 3 : MAP1LC3B knockout HepG2 untreated control cell lysate
Lane 4 : MAP1LC3B knockout HepG2 Treated Chloroquine (50 uM, 16 h) cell lysate
Lane 5 : U-87 MG cell lysate
Lane 6 : PC-12 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 15 kDa
Observed band size: 14,16 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-LC3B antibody [EPR18709] - Autophagosome Marker staining at 1/2000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab192890 was shown to bind specifically to LC3B. A band was observed at 16/14 kDa (yellow arrows) in treated wild-type HepG2 cell lysates with no signal observed at this size in MAP1LC3B knockout cell line ab277828 (knockout cell lysate ab283796). To generate this image, wild-type and MAP1LC3B knockout HepG2 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
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Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: LC3B knockout HAP1 cell lysate (20 µg)
Lane 3: Human brain tissue lysate (20 µg)
Lane 4: U-87 MG cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green).Green -target observed at 14 and 16 kDa. Red - loading control, ab8245, observed at 37 kDa.
This western blot image is a comparison between ab192890 and a competitor's top cited rabbit polyclonal antibody.
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ab192890 staining LC3B in HeLa (Human epithelial cell line from cervix adenocarcinoma) cells +/- Chloroquine (50μM, 24 hours).
The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab192890 at 1μg/ml and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 594), at 1/250 dilution (shown in pseudocolor red) followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labeled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
Alexa Fluor® 488 (ab225383) and Alexa Fluor® 647 (ab225382) conjugated versions are available for this clone.
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ab192890 staining LC3B in paraffin embedded human astrocytoma tissue by Immunohistochemistry. Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins. Samples were incubated with primary antibody at 1/1000 dilution for 30 mins at room temperature. Ready to use LeicaDS9800 (Bond™ Polymer Refine Detection) was used as the secondary antibody. Hematoxylin was used as a counterstain. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.
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Immunohistochemical analysis of formalin fixed paraffin embedded human cortex labelling LC3B with ab192890 at a dilution of 0.1 µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with an OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was conducted for 32min with ULTRA cell conditioning solution (CC1 pH8.5) . ab192890 anti LC3B antibody was incubated at 37°C for 16min. Sections were counterstained is with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control
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Tissue Microarrays stained for Anti-LC3B antibody [EPR18709] - Autophagosome Marker using ab192890 in immunohistochemical analysis. This table provides a detailed overview of positive (tick mark) and negaive (cross mark) staining per sample type tested.
Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.
The section was incubated with ab192890 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument. -
IHC image of LC3B staining in a section of formalin-fixed paraffin-embedded normal human cerebral cortex* performed on a Leica BONDTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20mins. The section was then incubated with ab221794, 0.1ug/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
This data was developed using the same antibody clone in a different buffer formulation containing only PBS (ab221794).
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ab192890 staining LC3B in HAP1 cells (wildtype and MAP1LC3B knockout) +/- Chloroquine (50μM, 24 hours).
The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab192890 at 1 μg/ml and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 594), at 1/250 dilution (shown in pseudocolor red) followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labeled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
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Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: LC3B knockout HAP1 cell lysate (20 µg)
Lane 3: Human brain tissue lysate (20 µg)
Lane 4: U-87 MG cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab192890 observed at 14 and 16 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab192890 was shown to specifically react with LC3B in wild-type HAP1 cells. No band was observed when LC3B knockout samples were examined. Wild-type and LC3B knockout samples were subjected to SDS-PAGE. ab192890 and ab8245 (loading control to GAPDH) were diluted 1/2000 and 1/10,000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging. -
All lanes : Anti-LC3B antibody [EPR18709] - Autophagosome Marker (ab192890) at 1/2000 dilution
Lane 1 : Human brain lysate
Lane 2 : U-87 MG (Human glioblastoma-astrocytoma epithelial cell line) whole cell lysate
Lane 3 : C6 (Rat glial tumor cell line) whole cell lysate
Lane 4 : RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate
Lane 5 : Mouse heart lysate
Lane 6 : Rat heart lysate
Lane 7 : Mouse brain lysate
Lane 8 : Rat brain lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/10000 dilution
Predicted band size: 15 kDa
Observed band size: 14,16 kDa why is the actual band size different from the predicted?Blocking/Dilution buffer: 5% NFDM/TBST.
Exposure times: Lane 1-6: 3 minutes; Lane 7 and 8: 30 seconds.
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Different batches of ab192890 were tested onU-87 MG (Human glioblastoma-astrocytoma epithelial cell line) lysate at 0.9 µg/ml. 15 µg of lysate was loaded in each lane. Bands observed at 14,16 kDa.
实验方案
数据表及文件
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SDS download
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Datasheet download
Certificate of Compliance
文献 (305)
ab192890 被引用在 305 文献中.
- Gao P et al. circEXOC5 promotes acute lung injury through the PTBP1/Skp2/Runx2 axis to activate autophagy. Life Sci Alliance 6:N/A (2023). PubMed: 36302650
- Wang W et al. Zinc-finger protein CXXC5 promotes breast carcinogenesis by regulating the TSC1/mTOR signaling pathway. J Biol Chem 299:102812 (2023). PubMed: 36539038
- Jiang B et al. Combination of chloroquine diphosphate and salidroside induces human liver cell apoptosis via regulation of mitochondrial dysfunction and autophagy. Mol Med Rep 27:N/A (2023). PubMed: 36579660
- Cui Y et al. Electroacupuncture attenuates spared nerve injury-induced neuropathic pain possibly by promoting the progression of AMPK/mTOR-mediated autophagy in spinal microglia. Ann Transl Med 10:1278 (2022). PubMed: 36618785
- Li P et al. 4-Hydroxyisoleucine inhibits tumor growth by triggering endoplasmic reticulum stress and autophagy. Curr Res Pharmacol Drug Discov 3:100127 (2022). PubMed: 36568272