Anti-160 kD Neurofilament Medium 抗体 [EPR23510-76] - BSA and Azide free (ab281830)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR23510-76] to 160 kD Neurofilament Medium - BSA and Azide free
- Suitable for: WB, IP, IHC-P, ELISA, IHC-Fr, Flow Cyt (Intra), ICC/IF
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
製品の概要
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製品名
Anti-160 kD Neurofilament Medium antibody [EPR23510-76] - BSA and Azide free
160 kD Neurofilament Medium 一次抗体 製品一覧 -
製品の詳細
Rabbit monoclonal [EPR23510-76] to 160 kD Neurofilament Medium - BSA and Azide free -
由来種
Rabbit -
アプリケーション
適用あり: WB, IP, IHC-P, ELISA, IHC-Fr, Flow Cyt (Intra), ICC/IFmore details -
種交差性
交差種: Mouse, Rat, Human -
免疫原
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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ポジティブ・コントロール
- WB: Human cerebellum and nerve tissue lysate. Mouse and rat brain and cerebellum tissue lysate. Wild-type HEK-293T lysate. IHC-P: Human cerebellum and cerebrum tissue. Mouse and rat cerebellum tissue. IHC-Fr: Mouse and rat cerebrum tissue. ICC/IF: Parental HEK-293T cells. Mouse and rat primary neural/glia cells. Flow Cyt: Wild-type HEK-293T cells. Mouse and rat primary neuron cells. IP: Mouse and rat whole cell lysate.
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特記事項
ab281830 is the carrier-free version of ab254348.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
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.
製品の特性
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製品の状態
Liquid -
保存方法
Shipped at 4°C. Store at +4°C. -
バッファー
Constituent: 100% PBS -
キャリア・フリー
はい -
Concentration information loading...
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精製度
Protein A purified -
ポリ/モノ
モノクローナル -
クローン名
EPR23510-76 -
アイソタイプ
IgG -
研究分野
関連製品
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Alternative Versions
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Compatible Secondaries
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Isotype control
アプリケーション
The Abpromise guarantee
Abpromise保証は、 次のテスト済みアプリケーションにおけるab281830の使用に適用されます
アプリケーションノートには、推奨の開始希釈率がありますが、適切な希釈率につきましてはご検討ください。
アプリケーション | Abreviews | 特記事項 |
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 102 kDa.
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IP |
Use at an assay dependent concentration.
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IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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ELISA |
Use at an assay dependent concentration.
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IHC-Fr |
Use at an assay dependent concentration.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20) |
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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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ICC/IF |
Use at an assay dependent concentration.
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特記事項 |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 102 kDa. |
IP
Use at an assay dependent concentration. |
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
ELISA
Use at an assay dependent concentration. |
IHC-Fr
Use at an assay dependent concentration. Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20) |
Flow Cyt (Intra)
Use at an assay dependent concentration. |
ICC/IF
Use at an assay dependent concentration. |
ターゲット情報
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機能
Neurofilaments usually contain three intermediate filament proteins: L, M, and H which are involved in the maintenance of neuronal caliber. -
配列類似性
Belongs to the intermediate filament family. -
翻訳後修飾
There are a number of repeats of the tripeptide K-S-P, NFM is phosphorylated on a number of the serines in this motif. It is thought that phosphorylation of NFM results in the formation of interfilament cross bridges that are important in the maintenance of axonal caliber.
Phosphorylation seems to play a major role in the functioning of the larger neurofilament polypeptides (NF-M and NF-H), the levels of phosphorylation being altered developmentally and coincidentally with a change in the neurofilament function.
Phosphorylated in the head and rod regions by the PKC kinase PKN1, leading to the inhibition of polymerization. - Information by UniProt
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参照データベース
- Entrez Gene: 4741 Human
- Entrez Gene: 18040 Mouse
- Entrez Gene: 24588 Rat
- Omim: 162250 Human
- SwissProt: P07197 Human
- SwissProt: P08553 Mouse
- SwissProt: P12839 Rat
- Unigene: 458657 Human
see all -
別名
- 150kDa medium antibody
- 160 kDa neurofilament protein antibody
- NEF3 antibody
see all
画像
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All lanes : Anti-160 kD Neurofilament Medium antibody [EPR23510-76] (ab254348) at 1/1000 dilution
Lane 1 : Human cerebellum tissue lysate
Lane 2 : Human nerve tissue lysate
Lane 3 : Human liver tissue lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/1000 dilution
Predicted band size: 102 kDa
Observed band size: 160 kDa why is the actual band size different from the predicted?This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Blocking and diluting buffer and concentration: 5% NFDM/TBST.
The molecular weight is consistent to what has been described in the literature (PMID:19239416, PMID:27000625).
Negative control: liver (PMID:30541916).
Exposure time: 10 secs.
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded human cerebellum tissue labelling 160 kD Neurofilament Medium with ab254348 at 1/4000 (0.111 µg/ml) dilution followed by ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). The section was incubated with ab254348 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Positive staining on human cerebellum.
Secondary antibody only control: Secondary antibody is ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded mouse liver tissue labelling 160 kD Neurofilament Medium with ab254348 at 1/4000 (0.111 µg/ml) dilution followed by ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). The section was incubated with ab254348 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Negative control: No staining on mouse liver.
Secondary antibody only control: Secondary antibody is ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunofluorescent analysis of parental HEK-293T (EDWT04) and NEFM KO HEK-293T (ED040746) cells labelling 160 kD Neurofilament Medium with ab254348 at 1/50 (8.86 µg/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The nuclear counterstain was (Blue).
Confocal image showing cytoplasmic staining in parental HEK-293T cells and no staining in NEFM KO HEK-293T cells.
Secondary antibody only control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunofluorescent analysis of mouse primary neural/glia cells fixed with 4% Paraformaldehyde and permeabilised with 0.1% TritonX-100, labelling 160 kD Neurofilament Medium with ab254348 at 1/50 (8.86 µg/ml) dilution. ab11267 Anti-MAP2 mouse monoclonal antibody was used to counterstain at 1/500 (4 μg/ml). Followed by secondary ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 (2 μg/ml) (Green). ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) was used as secondary counterstain at 1/1000 (2 μg/ml) (Red). The nuclear counterstain was DAPI (Blue).
Confocal image showing cytoplasmic staining in mouse primary neuron cells.
Confocal scanning Z step was set as 0.3 μm followed by image processing with maximum Z projection.Negative Control 1: ab150120 1/1000 (2 μg/ml)
Negative Control 2: ab11267 1/500 (4 μg/ml), secondary: ab150077 1/1000 (2μg/ml)
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunofluorescent analysis of rat primary neural/glia cells fixed with 4% Paraformaldehyde and permeabilised with 0.1% TritonX-100, labelling 160 kD Neurofilament Medium with ab254348 at 1/50 (8.86 µg/ml) dilution. ab11267 Anti-MAP2 mouse monoclonal antibody was used to counterstain at 1/500 (4 μg/ml). Followed by secondary ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 (2 μg/ml) (Green). ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) was used as secondary counterstain at 1/1000 (2 μg/ml) (Red). The nuclear counterstain was DAPI (Blue).
Confocal image showing cytoplasmic staining in rat primary neuron cells.
Confocal scanning Z step was set as 0.3 μm followed by image processing with maximum Z projection.Negative Control 1: ab150120 1/1000 (2 μg/ml)
Negative Control 2: ab11267 1/500 (4 μg/ml), secondary: ab150077 1/1000 (2 μg/ml)
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized wild-type HEK-293T (human embryonic kidney epithelial cell, Right)/ 160 kD Neurofilament Medium knockout HEK-293T cells (Left) labelling 160 kD Neurofilament Medium with ab254348 at 1/500 dilution (0.1 µg). A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
Positive staining on human wild-type HEK-293T cell line (ab255449), while no staining on human NEFM knockout HEK-293T cells (ab266741).
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse cerebrum tissue labeling 160 kD Neurofilament Medium with ab254348 at 1/100 (4.43 µg/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). The nuclear counterstain was DAPI (Blue).
Positive staining on mouse cerebrum is observed.
Secondary antibody control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488 )at 1/1000 dilution.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).
-
This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse liver tissue labeling 160 kD Neurofilament Medium with ab254348 at 1/100 (4.43 µg/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). The nuclear counterstain was DAPI (blue).
Negative control: No staining on mouse liver (PMID: 30541916) is observed.
Secondary antibody control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
160 kD Neurofilament Medium was immunoprecipitated from 0.35 mg mouse brain tissue lysate 10 µg with ab254348 at 1/30 dilution (2 µg in 0.35 mg lysates). Western blot was performed on the immunoprecipitate using ab254348 at 1/1000 dilution. VeriBlot for IP secondary antibody (HRP) (ab131366) was used at 1/5000 dilution.
Lane 1: Mouse brain tissue lysate 10 µg.
Lane 2: ab254348 IP in mouse brain tissue lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab254348 in mouse brain tissue lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 3.25 secs.
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
ELISA analysis using ab254348 at a range of 0-1000 ng/ml followed by a Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L)at 1/2500 dilution. Antigen concentration: 1000 ng/ml.
Antigens: Mouse 160 kD Neurofilament Medium.
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All lanes : Anti-160 kD Neurofilament Medium antibody [EPR23510-76] (ab254348) at 1/1000 dilution
Lane 1 : Mouse brain tissue lysate
Lane 2 : Mouse cerebellum tissue lysate
Lane 3 : Mouse liver tissue lysate
Lane 4 : Rat brain tissue lysate
Lane 5 : Rat cerebellum tissue lysate
Lane 6 : Rat liver tissue lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 102 kDa
Observed band size: 160 kDa why is the actual band size different from the predicted?This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Blocking and diluting buffer and concentration: 5% NFDM/TBST.
The molecular weight is consistent to what has been described in the literature (PMID:19239416, PMID:27000625.
Negative control: liver (PMID:30541916).
Exposure times: Lane 1-3: 3.25 secs; Lane 4-6: 10 secs.
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All lanes : Anti-160 kD Neurofilament Medium antibody [EPR23510-76] (ab254348) at 1/1000 dilution
Lane 1 : Wild-type HEK-293T (human embryonic kidney epithelial cell) whole cell lysate
Lane 2 : NEFM knockout HEK-293T whole cell lysate
Lane 3 : A549 (human lung carcinoma epithelial cell) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) at 1/10000 dilution
Predicted band size: 102 kDa
Observed band size: 160 kDa why is the actual band size different from the predicted?This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Blocking and diluting buffer and concentration: Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS.
Lanes 1-3: Merged signal (red and green). Green - ab254348 observed at 160 kDa. Red - loading control ab8245 observed at 36 kDa.
ab254348 Anti-NEFM antibody [EPR23510-76] was shown to specifically react with NEFM in wild-type HEK-293T cells. Loss of signal was observed when the knockout cell line ab266741 (knockout cell lysate ab257103) was used. Wild-type and NEFM knockout samples were subjected to SDS-PAGE. ab254348 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at at 4°C overnight at 1 in 1000 dilution and 1 in 20000 dilution respectively. 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 in 10000 dilution for 1 hour at room temperature before imaging.
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded human cerebrum tissue labelling 160 kD Neurofilament Medium with ab254348 at 1/4000 (0.111 µg/ml) dilution followed by ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). The section was incubated with ab254348 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Positive staining on human cerebrum.
Secondary antibody only control: Secondary antibody is ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
-
This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded mouse cerebellum tissue labelling 160 kD Neurofilament Medium with ab254348 at 1/4000 (0.111 µg/ml) dilution followed by ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). The section was incubated with ab254348 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Positive staining on mouse cerebellum.
Secondary antibody only control: Secondary antibody is ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
-
This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded rat cerebellum tissue labelling 160 kD Neurofilament Medium with ab254348 at 1/4000 (0.111 µg/ml) dilution followed by ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). The section was incubated with ab254348 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Positive staining on rat cerebellum.
Secondary antibody only control: Secondary antibody is ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
-
This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat cerebrum tissue labeling 160 kD Neurofilament Medium with ab254348 at 1/100 (4.43 µg/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). The nuclear counterstain was DAPI (Blue).
Positive staining on rat cerebrum is observed.
Secondary antibody control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488 )at 1/1000 dilution.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).
-
This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat liver tissue labeling 160 kD Neurofilament Medium with ab254348 at 1/100 (4.43 µg/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). The nuclear counterstain was DAPI (blue).
Negative control: No staining on rat liver (PMID: 30541916) is observed.
Secondary antibody control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).
-
This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Flow cytometric analysis of 4% paraformaldehyde fixed, 90% methanol permeabilized mouse primary neuron cells labelling 160 kD Neurofilament Medium with ab254348 at 1/500 dilution (0.1 µg)/ Right compared with a Rabbit monoclonal IgG isotype control (ab172730) / Left.
A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
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This data was developed using ab254348, the same antibody clone in a different buffer formulation.
Flow cytometric analysis of 4% paraformaldehyde fixed, 90% methanol permeabilized rat primary neuron cells cells labelling 160 kD Neurofilament Medium with ab254348 at 1/500 dilution (0.1 µg)/ Right compared with a Rabbit monoclonal IgG isotype control (ab172730) / Left.
A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
-
This data was developed using ab254348, the same antibody clone in a different buffer formulation.
160 kD Neurofilament Medium was immunoprecipitated from 0.35 mg rat brain tissue lysate 10 µg with ab254348 at 1/30 dilution (2 µg in 0.35 mg lysates). Western blot was performed on the immunoprecipitate using ab254348 at 1/1000 dilution. VeriBlot for IP secondary antibody (HRP) (ab131366) was used at 1/5000 dilution.
Lane 1: Rat brain tissue lysate 10 µg.
Lane 2: ab254348 IP in rat brain tissue lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab254348 in rat brain tissue lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 5.5 secs.
プロトコール
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
データシートおよび資料
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Datasheet download
Certificate of Compliance
参考文献 (0)
ab281830 は論文での使用が確認できていません。