製品の概要

  • 製品名
    Anti-LAMP1 antibody [H4A3]
    LAMP1 一次抗体 製品一覧
  • 製品の詳細
    Mouse monoclonal [H4A3] to LAMP1
  • 由来種
    Mouse
  • アプリケーション
    適用あり: ICC/IF, Flow Cyt, IHC-Fr, WB, IHC-Pmore details
  • 種交差性
    交差種: Rat, Human, Monkey
  • 免疫原

    The details of the immunogen for this antibody are not available.

  • ポジティブ・コントロール
    • ICC/IF: Hela cells; Human gastric cancer cells; HaCaT keratinocytes. IHC-P: Human placenta tissue. WB: Jurkat and MCF-7 cells.
  • 特記事項

    Although there are publications stating this antibody works with Mouse species (PMID 18840681), and previous batches gave positive staining on murine cells, recent batches are failing to react with this species (from customer feedback). Further feedback using this antibody with Mouse tissues would be very welcome.

    This product was changed from ascites to tissue culture supernatant on 3rd September 2018. Please note that the dilutions may need to be adjusted accordingly. If you have any questions, please do not hesitate to contact our scientific support team.

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab25630 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

アプリケーション Abreviews 特記事項
ICC/IF Use a concentration of 5 - 10 µg/ml.
Flow Cyt 1/20.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

IHC-Fr Use a concentration of 1 µg/ml.
WB 1/10000. Detects a band of approximately 48 kDa (predicted molecular weight: 45 kDa).
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

ターゲット情報

  • 機能
    Presents carbohydrate ligands to selectins. Also implicated in tumor cell metastasis.
  • 配列類似性
    Belongs to the LAMP family.
  • 翻訳後修飾
    O- and N-glycosylated; some of the 18 N-linked glycans are polylactosaminoglycans.
  • 細胞内局在
    Cell membrane. Endosome membrane. Lysosome membrane. This protein shuttles between lysosomes, endosomes, and the plasma membrane.
  • Information by UniProt
  • 参照データベース
  • 別名
    • CD107 antigen like family member A antibody
    • CD107 antigen-like family member A antibody
    • CD107a antibody
    • CD107a antigen antibody
    • LAMP 1 antibody
    • LAMP-1 antibody
    • LAMP1 antibody
    • LAMP1_HUMAN antibody
    • LAMPA antibody
    • LGP120 antibody
    • lgpA antibody
    • Lysosomal membrane glycoprotein 120KD antibody
    • Lysosomal Associated Membrane Protein 1 antibody
    • Lysosome associated membrane glycoprotein 1 antibody
    • Lysosome-associated membrane glycoprotein 1 antibody
    • Lysosome-associated membrane protein 1 antibody
    • OTTHUMP00000040663 antibody
    see all

画像

  • Ab25630 stained Hela cells. The cells were 100% methanol fixed for 5 minutes and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Triton for 1h at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with ab25630 at 5µg/ml and ab202272 (Rabbit monoclonal [EP1332Y] to alpha Tubulin - Microtubule Marker (Alexa Fluor® 594) – pseudo-colored red) overnight at +4°C. The secondary antibody (pseudo-colored green) was a Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed (ab150117) used at a 1/1000 dilution for 1h at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43µM for 1hour at room temperature.

  • IHC image of LAMP1 staining in human placenta formalin fixed paraffin embedded tissue section*, performed on a Leica Bond™ 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 20 mins. The section was then incubated with ab25630, 1µg/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.

    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.

    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

  • All lanes : Anti-LAMP1 antibody [H4A3] (ab25630) at 1/1000 dilution

    Lane 1 : Jurkat cell lysate
    Lane 2 : Jurkat membrane lysate
    Lane 3 : MCF-7 cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 45 kDa
    Observed band size: 115-120 kDa
    why is the actual band size different from the predicted?



    This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using Licor blocking buffer before being incubated with ab25630 (1/1000) overnight at 4°C. Antibody binding was detected using ab9485 (rabbit anti-GAPDH);  at a 1/10,000 dilution for 1hr at room temperature and then imaged using the Licor Odyssey CLx.

  • Overlay histogram showing Jurkat cells stained with ab25630 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab25630, 1/20 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in Jurkat cells fixed with methanol (5 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

  • ab25630 at 1/500 dilution staining LAMP1 in human gastric cancer cells by immunocytochemistry/ immunofluorescence. Sections were methanol fixed, permeabilized in 0.5% Triton X-100 prior to blocking in 10% NGS/1% BSA for 1 hour at 25°C and then incubated with ab25630 for 2 hours at 25°C. Alexa fluor® 594 mouse polyclonal to mouse Ig, diluted 1/600, was used as the secondary antibody.

    See Abreview

  • ab25630 at 1/100 staining human Primary Gingival epithelial cells by ICC/IF. The cells were ixed with 4% paraformaldehyde, permeabilized with 0.5% saponin and then blocked overnight with 10% goat serum, 5% BSA. The cells were incubated with the antibody for 1 hour and then a FITC conjugated goat polyclonal antibody was used as the secondary. The cells were counterstaind with DAPI for the nucleus and Cell Tracker Blue for the cytoplasm.

    See Abreview

  • Ab25630 staining Human normal placenta. Staining is localized to the cell membrane.
    Left panel: with primary antibody at 1 ug/ml. Right panel: isotype control.
    Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 AR buffers EDTA pH 9.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.
  • ab25630 staining LAMP1 in human HaCaT keratinocytes by Immunocytochemistry/ Immunofluorescence. Cells were fixed with acetone, permeabilized with ice cold acetone and blocking with 4% PBS, 0.4% BSA and goat serum was performed for 16 hours at 40C. Samples were incubated with primary antibody (1/100: in 10% blocking solution in PBS) for 1 hour at 230C. An Alexa Fluor ® 680-conjugated goat polyclonal to mouse IgG was used at dilution at 1/200 as secondary antibody. Alexafluor-680 signal is pseudocolored green in the image.

    See Abreview

  • IHC-Fr image of human brain section stained with ab25630. The brain sections were incubated in 10% normal donkey serum in 0.1% PBS- and 0.3x triton100 for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab25630, 1µg/ml) overnight at +4°C. The secondary antibody was Alexa Fluor®568 donkey anti-mouse IgG (H+L) used at a 1/1000 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

    See Abreview

  • All lanes : Anti-LAMP1 antibody [H4A3] (ab25630) at 1/10000 dilution

    Lane 1 : Iron treated 3 month old liver at 20 µg
    Lane 2 : Untreated 3 month old liver at 20 µg
    Lane 3 : One month old untreated liver

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 45 kDa
    Observed band size: 120,48 kDa why is the actual band size different from the predicted?
    Additional bands at: 120 kDa (possible glycosylated form)


    Exposure time: 5 minutes


    WB image of LAMP1 (ab25630) on Mouse liver. Lanes were loaded 20 ug of Liver tissue lysate Lane 1. iron treated 3 month old liver, lane 2. untreated 3 month old liver, Lane 3. one month old untreated liver.

    See Abreview

参考文献

This product has been referenced in:
  • Nahorski MS  et al. Clathrin heavy chain 22 contributes to the control of neuropeptide degradation and secretion during neuronal development. Sci Rep 8:2340 (2018). Read more (PubMed: 29402896) »
  • Sørensen DM  et al. Parkinson disease related ATP13A2 evolved early in animal evolution. PLoS One 13:e0193228 (2018). Read more (PubMed: 29505581) »
See all 102 Publications for this product

レビューと Q&A

1-2 of 2 Q&A

Question
Answer

The predicted calculated MW for LAMP1 is approximately 44.8 kDa (human) and 43.9 kDa (rat). However, the observed MW for LAMP1 as run on an SDS-PAGE gel can be much higher, and the protein is heavily glycosylated in both species. As such, the predicted observed MW for LAMP1 in both human and rat is anticipated to be approximately in the range of 110-140 kDa.

Additional information on LAMP1 and its post-translational modifications can be found here:

UniProt Human LAMP1 P11279

UniProt Rat LAMP1 P14562

Read More

Question
Answer

Yes – we do have a conjugated format of anti-LAMP1 Ab clone 4HA3. This is an Alexa Fluor 488 conjugated version of the clone, and is referenced as product Anti-LAMP1 antibody [H4A3] (Alexa Fluor® 488) (ab187591). Please refer to the following product webpage for additional information:

Anti-LAMP1 antibody [H4A3] (Alexa Fluor® 488) (ab187591)

Read More

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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