Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)

Rabbit Recombinant Monoclonal LC3B antibody. Carrier free. Suitable for ICC/IF, WB, IHC-P and reacts with Human, Mouse, Rat samples. Cited in 10 publications.

Images

Immunocytochemistry/ Immunofluorescence - Anti-LC3B antibody [EPR18709] - BSA and Azide free (AB221794), expandable thumbnail

Publications

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Constituents: PBS
Form: Liquid
Clonality: Monoclonal

Immunogen

The exact immunogen used to generate this antibody is proprietary information.

Reactivity data

Select an application

Product promiseTestedExpectedPredictedNot recommended

	ICC/IF	IP	WB	IHC-P
Human	Tested	Not recommended	Tested	Tested
Mouse	Expected	Not recommended	Tested	Expected
Rat	Expected	Not recommended	Tested	Expected

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Expected
Expected

Species	Dilution info	Notes
Species Mouse, Rat	Dilution info Use at an assay dependent concentration.	Notes -

Not recommended
Not recommended

Species	Dilution info	Notes
Species Human, Mouse, Rat	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse, Rat, Human	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes Perform heat-mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

Expected
Expected

Species	Dilution info	Notes
Species Mouse, Rat	Dilution info Use at an assay dependent concentration.	Notes -

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Target data

See full target information MAP1LC3B

Function

Ubiquitin-like modifier involved in formation of autophagosomal vacuoles (autophagosomes) (PubMed:20418806, PubMed:23209295, PubMed:28017329). Plays a role in mitophagy which contributes to regulate mitochondrial quantity and quality by eliminating the mitochondria to a basal level to fulfill cellular energy requirements and preventing excess ROS production (PubMed:23209295, PubMed:28017329). In response to cellular stress and upon mitochondria fission, binds C-18 ceramides and anchors autophagolysosomes to outer mitochondrial membranes to eliminate damaged mitochondria (PubMed:22922758). While LC3s are involved in elongation of the phagophore membrane, the GABARAP/GATE-16 subfamily is essential for a later stage in autophagosome maturation (PubMed:20418806, PubMed:23209295, PubMed:28017329). Promotes primary ciliogenesis by removing OFD1 from centriolar satellites via the autophagic pathway (PubMed:24089205). Through its interaction with the reticulophagy receptor TEX264, participates in the remodeling of subdomains of the endoplasmic reticulum into autophagosomes upon nutrient stress, which then fuse with lysosomes for endoplasmic reticulum turnover (PubMed:31006537, PubMed:31006538). Upon nutrient stress, directly recruits cofactor JMY to the phagophore membrane surfaces and promotes JMY's actin nucleation activity and autophagosome biogenesis during autophagy (PubMed:30420355).

Alternative names

MAP1ALC3, MAP1LC3B, Microtubule-associated proteins 1A/1B light chain 3B, Autophagy-related protein LC3 B, Autophagy-related ubiquitin-like modifier LC3 B, MAP1 light chain 3-like protein 2, MAP1A/MAP1B light chain 3 B, Microtubule-associated protein 1 light chain 3 beta, MAP1A/MAP1B LC3 B

Recommended products

Rabbit Recombinant Monoclonal LC3B antibody. Carrier free. Suitable for ICC/IF, WB, IHC-P and reacts with Human, Mouse, Rat samples. Cited in 10 publications.

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Constituents: PBS
Form: Liquid
Clonality: Monoclonal
Immunogen: The exact immunogen used to generate this antibody is proprietary information.
Carrier free: Yes
Clone number: EPR18709
Purification technique: Affinity purification Protein A
Specificity: FURTHER INFORMATION ON SPECIFICITY (Chinese Version) available under the support & downloads section.
Concentration

Storage

Shipped at conditions: Blue Ice
Appropriate short-term storage conditions: +4°C
Appropriate long-term storage conditions: +4°C
Storage information: Do Not Freeze

Notes

ab221794 is the carrier-free version of Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890.

Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

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 batch production

For more information, read more on recombinant antibodies.

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.

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.

Activity summary

LC3B also known as microtubule-associated protein 1 light chain 3 beta plays an important role in autophagy a process that maintains cellular homeostasis. The LC3B protein with a molecular weight of approximately 14 kDa undergoes lipidation to form LC3-II which associates with autophagosomal membranes. This protein expresses ubiquitously in various tissues including liver muscle and brain. Researchers often detect LC3B using techniques like LC3B western blot and LC3B immunofluorescence due to its function as a marker indicating autophagy levels.

Biological function summary

LC3B contributes significantly to the formation and maturation of autophagosomes. LC3B part of the autophagy-related protein complex binds to autophagic membranes. During this process LC3-I converts to LC3-II a lipid-phosphatidylethanolamine conjugate essential for autophagosome membrane expansion and closure. This mechanism helps remove damaged organelles and misfolded proteins from cells therefore contributing to cellular quality control.

Pathways

LC3B integrates into the autophagy pathway which is critical for cellular adaptive responses to stress. The mammalian target of rapamycin (mTOR) pathway regulates autophagy where mTOR inhibition activates LC3B promoting autophagosome formation. Moreover LC3B operates alongside other proteins like Beclin-1 and ULK1 facilitating the initiation and progression of autophagy under nutrient starvation conditions. These interactions highlight LC3's role in cellular energy balance and survival mechanisms.

Associated diseases and disorders

LC3B connects with conditions such as cancer and neurodegenerative diseases. Altered autophagy levels mediated by LC3B often associate with tumorigenesis where its dysregulation can affect cancer progression. Furthermore LC3B also links to neurodegenerative diseases like Alzheimer's where impaired autophagy disrupts neuronal function. LC3B interacts with proteins such as p62/SQSTM1 which affects protein aggregate clearance a critical factor in neurodegenerative pathology.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
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11 product images

Immunocytochemistry/ Immunofluorescence - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
LC3B immunofluorescence staining of HeLa cells using rabbit anti-LC3B antibody

Anti-LC3B antibody [EPR18709] - Autophagosome Marker 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 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890 at 1μg/ml and Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker 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) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed 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 (Alexa Fluor® 647 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab225383) and Alexa Fluor^® 647 (Alexa Fluor® 488 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab225382) conjugated versions are available for this clone.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890).
Immunoprecipitation - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
LC3B was immunoprecipitated from 1mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890 at 1/50 dilution. Western blot was performed from the immunoprecipitate using Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890 at 1/1000 dilution. Veriblot for IP (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366), was used for detection at 1/1000 dilution.
Lane 1: HeLa whole cell lysate 10μg (Input).
Lane 2: Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890 IP in Jurkat whole cell lysate.
Lane 3: Rabbit IgG,monoclonal [EPR25A] -Isotype Control (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890 in HeLa whole cell cell lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 30 seconds.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890).
All lanes: Immunoprecipitation - Anti-LC3B antibody [EPR18709] - Autophagosome Marker (Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890)
Predicted band size: 15 kDa
Exposure time: 30s
Immunocytochemistry/ Immunofluorescence - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
LC3B immunofluorescence staining of HAP1 cells using rabbit anti-LC3B antibody

This ICC data was generated using the same anti-LC3B antibody clone, EPR18709, in a different buffer formulation (cat# Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890).
Anti-LC3B antibody [EPR18709] - Autophagosome Marker 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 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890 at 1 μg/ml and Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker 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) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed 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).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
IHC image of LC3B staining in a section of formalin-fixed paraffin-embedded normal human cerebral cortex* performed on a Leica BOND^TM 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.
Immunocytochemistry/ Immunofluorescence - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
Clone EPR18709 (ab221794) has been successfully conjugated by Abcam. This image was generated using Anti-LC3B antibody [EPR18709] - Autophagosome Marker (Alexa Fluor® 647). Please refer to Alexa Fluor® 647 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab225383 for protocol details.
Alexa Fluor® 647 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab225383 staining LC3B in wild-type HAP1 cells and knockout cells, untreated and chloroquine-treated (50μM, 24 hours). The cells were fixed with 100% methanol (5min), 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 with Alexa Fluor® 647 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab225383 at 0.5μg/ml (shown in red) and Alexa Fluor® 488 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195887, Mouse monoclonal to alpha Tubulin (Alexa Fluor^® 488), at 1/250 dilution (shown in green) overnight at +4°C. Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
Immunocytochemistry/ Immunofluorescence - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
Clone EPR18709 (ab221794) has been successfully conjugated by Abcam. This image was generated using Anti-LC3B antibody [EPR18709] - Autophagosome Marker (Alexa Fluor® 488). Please refer to Alexa Fluor® 488 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab225382 for protocol details.
Alexa Fluor® 488 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab225382 staining LC3B in HeLa chloroquine-treated (50µM, 24 hours) cells. 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 Alexa Fluor® 647 Anti-SQSTM1 / p62 antibody [EPR18351] ab225453 at 1/100 dilution (shown in green) and Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor^® 594), at 1/250 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
Western blot - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
This data was developed using Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890, the same antibody clone in a different buffer formulation. Different batches of Anti-LC3B antibody [EPR18709] - Autophagosome Marker 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.
All lanes: Western blot - Anti-LC3B antibody [EPR18709] - Autophagosome Marker (Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890)
Predicted band size: 15 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890).

Tissue Microarrays stained for Anti-LC3B antibody [EPR18709] - Autophagosome Marker using Anti-LC3B antibody [EPR18709] - Autophagosome Marker 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 Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
LC3B immunohistochemistry staining of human cortex using rabbit anti-LC3B antibody

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890).
Immunohistochemical analysis of formalin fixed paraffin embedded human cortex labelling LC3B with Anti-LC3B antibody [EPR18709] - Autophagosome Marker 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) . Anti-LC3B antibody [EPR18709] - Autophagosome Marker 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.
Western blot - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890).
False colour image of Western blot: Anti-LC3B antibody staining at 1 ug/ml, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-LC3B antibody ab63817 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.
All lanes: Western blot - Anti-LC3B antibody [EPR18709] - Autophagosome Marker (Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890) at 1/1000 dilution
Lane 1: Wild-type HepG2 untreated control cell lysate at 20 µg
Lane 2: Wild-type HepG2 Treated Chloroquine (50 uM, 16 h) cell lysate at 20 µg
Lane 2: Western blot - Human MAP1LC3B knockout Hep G2 cell line (ab277828)
Lane 2: Western blot - Human MAP1LC3B knockout Hep G2 cell lysate (ab283796)
Lane 3: MAP1LC3B knockout HepG2 untreated control cell lysate at 20 µg
Lane 4: MAP1LC3B knockout HepG2 Treated Chloroquine (50 uM, 16 h) cell lysate at 20 µg
Lane 5: U-87 MG cell lysate at 20 µg
Lane 6: PC-12 cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 15 kDa
Observed band size: 14 kDa, 16 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LC3B antibody [EPR18709] - BSA and Azide free (ab221794)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-LC3B antibody [EPR18709] - Autophagosome Marker ab192890).
Anti-LC3B antibody [EPR18709] - Autophagosome Marker 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.