Anti-LC3B antibody - Autophagosome Marker

9 product images

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  Portovedo M. et al PLoS One. 2015 Mar 18;10(3):e0119850. doi: 10.1371/journal.pone.0119850. eCollection 2015 Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  Immunostaining of LC3 (green) in the arcuate hypothalamic nucleus of lean and obese mice (A; 8 weeks of a HFD or B; 16 weeks of a HFD). DAPI (blue) was used for nuclear staining.
  

  The brain was excised after the mice were decapitated. Each SNC was fixed in 4% paraformaldehyde and each hypothalamus was processed for paraffin embedding and sectioned into 5.0 μm sections. Samples were incubated with primary antibodies overnight and with secondary antibodies conjugated to FITC or rhodamine for 2 hours (sc2777and sc2092, respectively; Santa Cruz Biotechnology, Santa Cruz, CA). The DAPI stain was used for nuclear staining while the Leica FW 4500 B microscope captured the images. Hypothalamic areas were observed according to the landmarks in the mouse brain atlas. Analysis and documentation of the results were performed using Leica Application Suite V3.6 (Switzerland).

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  Western blot - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  Western Blot shows lysates of HeLa (human epithelial cell line from cervix adenocarcinoma) cell line and LC3B knockout HeLa cell line (KO) untreated (-) or treated (+) with 50 uM Chloroquinine for 18 hours. PVDF membrane was probed with 0.5 ug/mL ab48394 followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody. A specific band was detected for LC3B at approximately 15 kDa (as indicated) in the parental HeLa cell line, but is not detectable in the knockout HeLa cell line. GAPDH is shown as a loading control. This experiment was conducted under reducing conditions.

  All lanes: Western blot - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  Predicted band size: 15 kDa

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  Immunocytochemistry/ Immunofluorescence - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  HeLa (human epithelial cell line from cervix adenocarcinoma) cells (wild type, left; LC3B knockout HeLa, right) stained for LC3B using ab48394 (red) at 0.3 μg/ml in ICC/IF. Primary antibody was incubated for 3 hours at room temperature, followed by NorthernLights™ 557-conjugated Anti-Rabbit IgG Secondary Antibody. Counterstained with DAPI (blue).
  

  LC3 was detected in immersion fixed Cloroquine treated Hela cells (left) but was not detected in LC3 knockout Hela cells (right).

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  Jiang C. et al PLoS Genet. 2017 Jan 10;13(1):e1006535. doi: 10.1371/journal.pgen.1006535. eCollection 2017 Jan. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  LC3B immunofluorescence in primary follicles of PD 13 ovaries. The red dots represent LC3b and DAPI (blue) indicated cell nuclei.
  

  For the immunofluorescence analysis, the ovaries were fixed in 4% paraformaldehyde, embedded in paraffin, and sectioned into 5 μm slices. After antigen retrieval, the slides were blocked with goat serum and incubated with primary antibody (rabbit anti-LC3B at 1:200) overnight at 4°C. Alexa Fluor 594 (Invitrogen) was used as the secondary antibody in immunofluorescence assays.

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  Formalin-fixed, paraffin-embedded mouse brain tissue stained for LC3B using ab48394 at 1/200 dilution in immunohistochemical analysis. The specific signal of LC3 was detected using HRP-conjugated secondary antibody with DAB reagent, and nuclei of cells were counterstained using hematoxylin. This LC3 antibody generated a low to moderate levels of cytoplasmic staining in the glial cells. The neurons depicted a moderate to strong staining for LC3 in their cytoplasm.

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  Western blot - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  Western blot shows lysates of mouse NIH/3T3 (mouse embryo fibroblast cell line) and rat PC-12 (rat adrenal gland pheochromocytoma cell line) cell lines untreated (-) or treated (+) with Chloroquine. PVDF membrane was probed with 0.5 ug/mL rabbit anti-LC3B polyclonal Antibody (ab48394), followed by 1:2000 dilution of goat anti-rabbit IgG secondary antibody.

  All lanes: Western blot - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  Predicted band size: 15 kDa

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  This image is courtesy of a customer review by Armen Petrosyan.

  Immunocytochemistry/ Immunofluorescence - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  ab48394 staining LC3B in a Rat hepatocyte by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with formaldehyde, permeabilized with 0.2% Triton X-100 in PBS and blocked with 1% Donkey serum in 0.1% PBST for 60 minutes at 21°C. Samples were incubated with primary antibody (1/50 in PBS + 1% BSA) for 3 hours at 22°C. An Alexa Fluor^® 394-conjugated Donkey anti-rabbit IgG polyclonal was used as the secondary antibody at a dilution of 1/200.

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  Immunocytochemistry/ Immunofluorescence - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  ab48394 staining LC3B in HeLa cells treated with calmidazolium chloride (Calmidazolium chloride, Calmodulin inhibitor ab120658), by ICC/IF. Increase of LC3B expression correlates with increased concentration of calmidazolium chloride, as described in literature.
  The cells were incubated at 37°C for 6h in media containing different concentrations of Calmidazolium chloride, Calmodulin inhibitor ab120658 (calmidazolium chloride) in DMSO, fixed with 4% formaldehyde for 10 minutes at room temperature and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab48394 (1 μg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 anti-rabbit polyclonal antibody (Goat Anti-Rabbit IgG H&L (DyLight® 488) preadsorbed ab96899) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.

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  This image is courtesy of an anonymous customer review

  Western blot - Anti-LC3B antibody - Autophagosome Marker (ab48394)

  All lanes: Western blot - Anti-LC3B antibody - Autophagosome Marker (ab48394) at 1/2000 dilution

  Lane 1: Rat whole tissue lysate - Normal liver at 30 µg

  Lane 2: Rat whole tissue lysate - liver treated with AEE788 at 50 mg/kg 3 times a week for 1 week at 30 µg

  Lane 3: Rat whole tissue lysate - liver treated with RAD at 2.5 mg/kg daily for 1 week at 30 µg

  Secondary

  All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab6721) at 1/5000 dilution

  Developed using the ECL technique.

  Predicted band size: 15 kDa

  Observed band size: 17 kDa

  Exposure time: 1min