Synthetic peptide corresponding to Human LAMP2B aa 350 to the C-terminus (C terminal) conjugated to Keyhole Limpet Haemocyanin (KLH). LAMP2 has 3 distinct isoforms, LAMP2A, 2B & 2C which are expressed in different tissues and differ in sequence at the extreme C-terminus. Database link: P13473 (Peptide available as ab23321)
This antibody gave a positive signal in the following whole cell lysates: HeLa, HEK293.
This antibody gave a positive signal in the following Methanol fixed cell lines: HeLa.
It also gave a positive signal in Human skin tissue sections.
保存方法Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml.
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Use a concentration of 1 µg/ml. Detects a band of approximately 110 kDa (predicted molecular weight: 110 kDa).
Abcam recommends using milk as the blocking agent.
機能Implicated in tumor cell metastasis. May function in protection of the lysosomal membrane from autodigestion, maintenance of the acidic environment of the lysosome, adhesion when expressed on the cell surface (plasma membrane), and inter- and intracellular signal transduction. Protects cells from the toxic effects of methylating mutagens.
組織特異性Isoform LAMP-2A is highly expressed in placenta, lung and liver, less in kidney and pancreas, low in brain and skeletal muscle. Isoform LAMP-2B is highly expressed in skeletal muscle, less in brain, placenta, lung, kidney and pancreas, very low in liver.
配列類似性Belongs to the LAMP family.
翻訳後修飾O- and N-glycosylated; some of the 16 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.
Western blot - Anti-LAMP2B antibody - Lysosome Marker (ab18529)
All lanes : Anti-LAMP2B antibody - Lysosome Marker (ab18529) at 1 µg/ml
Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate Lane 2 : HEK293 (Human) Whole Cell Lysate
Lysates/proteins at 20 µg per lane.
Secondary Goat Anti-Rabbit IgG H&L (HRP) preadsorbed at 1/50000 dilution developed using the ECL technique
Performed under reducing conditions.
Predicted band size : 110 kDa Observed band size : 110 kDa Additional bands at : 42 kDa. We are unsure as to the identity of these extra bands.
Exposure time : 4 minutes
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 3% Milk before being incubated with ab18529 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406.
LAMP2B undergoes extensive glycosylation so ab18529 detects a band of 110 kDa by Western blot.
ICC/IF image of ab18529 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab18529, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).
IHC image of ab18529 staining in human skin formalin fixed paraffin embedded tissue section, 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 20 mins. The section was then incubated with ab18529, 5µg/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.