- PA1-46286 - Invitrogen Antibodies MAP1LC3B antibody

Submitted references Abnormal lipid metabolism in epidermal Langerhans cells mediates psoriasis-like dermatitis.

Zhang X, Li X, Wang Y, Chen Y, Hu Y, Guo C, Yu Z, Xu P, Ding Y, Mi QS, Wu J, Gu J, Shi Y
JCI insight 2022 Jul 8;7(13)

Inhibition of the C-X-C Motif Chemokine 12 (CXCL12) and Its Receptor CXCR4 Reduces Utero-Placental Expression of the VEGF System and Increases Utero-Placental Autophagy.

Ashley RL, Runyan CL, Maestas MM, Trigo E, Silver G
Frontiers in veterinary science 2021;8:650687

ATF4 links ER stress with reticulophagy in glioblastoma cells.

Zielke S, Kardo S, Zein L, Mari M, Covarrubias-Pinto A, Kinzler MN, Meyer N, Stolz A, Fulda S, Reggiori F, Kögel D, van Wijk S
Autophagy 2021 Sep;17(9):2432-2448

Ring finger protein 213 assembles into a sensor for ISGylated proteins with antimicrobial activity.

Thery F, Martina L, Asselman C, Zhang Y, Vessely M, Repo H, Sedeyn K, Moschonas GD, Bredow C, Teo QW, Zhang J, Leandro K, Eggermont D, De Sutter D, Boucher K, Hochepied T, Festjens N, Callewaert N, Saelens X, Dermaut B, Knobeloch KP, Beling A, Sanyal S, Radoshevich L, Eyckerman S, Impens F
Nature communications 2021 Oct 1;12(1):5772

The Biochemical Pathways of Nicotinamide-Derived Pyridones.

Hayat F, Sonavane M, Makarov MV, Trammell SAJ, McPherson P, Gassman NR, Migaud ME
International journal of molecular sciences 2021 Jan 24;22(3)

Long Term Pharmacological Perturbation of Autophagy in Mice: Are HCQ Injections a Relevant Choice?

Masson JD, Blanchet B, Periou B, Authier FJ, Mograbi B, Gherardi RK, Crépeaux G
Biomedicines 2020 Mar 1;8(3)

STF-62247 and pimozide induce autophagy and autophagic cell death in mouse embryonic fibroblasts.

Kinzler MN, Zielke S, Kardo S, Meyer N, Kögel D, van Wijk SJL, Fulda S
Scientific reports 2020 Jan 20;10(1):687

Lidocaine induces protective autophagy in rat C6 glioma cell line.

Izdebska M, Hałas-Wiśniewska M, Zielińska W, Klimaszewska-Wiśniewska A, Grzanka D, Gagat M
International journal of oncology 2019 Mar;54(3):1099-1111

CXCR4 signaling at the ovine fetal-maternal interface regulates vascularization, CD34+ cell presence, and autophagy in the endometrium†.

Runyan CL, McIntosh SZ, Maestas MM, Quinn KE, Boren BP, Ashley RL
Biology of reproduction 2019 Jul 1;101(1):102-111

Loperamide, pimozide, and STF-62247 trigger autophagy-dependent cell death in glioblastoma cells.

Zielke S, Meyer N, Mari M, Abou-El-Ardat K, Reggiori F, van Wijk SJL, Kögel D, Fulda S
Cell death & disease 2018 Sep 24;9(10):994

HSPB8 and the Cochaperone BAG3 Are Highly Expressed During the Synthetic Phase of Rat Myometrium Programming During Pregnancy.

Marsh NM, Wareham A, White BG, Miskiewicz EI, Landry J, MacPhee DJ
Biology of reproduction 2015 May;92(5):131

Autophagy in zebrafish.

Varga M, Fodor E, Vellai T
Methods (San Diego, Calif.) 2015 Mar;75:172-80

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot detection of LC3B in treated U87-MG (human glioblastoma astrocytoma) lysates using Product # PA1-46286.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of LC3B in U87-MG (human glioblastoma astrocytoma) lysates. Sample was incubated in LC3B polyclonal antibody (Product # PA1-46286).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of LC3B in 10 µg Human cervical carcinoma (HeLa) and Mouse Neuroblast cells (Neuro2a). Samples were incubated in LC3B polyclonal antibody (Product # PA1-46286 using a dilution of 2 µg/mL. Treated with (+) and without (-) 50 µM Chloroquine overnight. Whole cell protein lysates were prepared in 1x Laemmli sample buffer and each lysate was separated on a 4–20% gel by SDS-PAGE, transferred to 0.2 µm PVDF membrane and blocked in 5% nonfat milk in TBST. Loading control: 1 µg/mL anti-alpha tubulin as a loading control. Detection: chemiluminescence substrate.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of LC3B in 0.5 mg/mL Neuro2A lysate. Samples were incubated in LC3B polyclonal (Product # PA1-46286). This experiment was performed under reducing conditions using the 12-230 kDa separation system.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Staining of treated U373-MG (human glioblastoma) cells using Product # PA1-46286.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunocytochemistry analysis of LC3B in treated U373-MG cells. Samples were incubated in LC3B polyclonal antibody (Product # PA1-46286). Analysis using the HRP conjugate of anti-LC3B . The nuclei were stained with DAPI.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunocytochemistry analysis of LC3B in HeLa cells. Samples were incubated in LC3B polyclonal antibody (Product # PA1-46286) followed by Alexa Fluor 488-conjugated Goat to rabbit IgG secondary antibody (green). Actin filaments were labeled with Alexa Fluor 568 phalloidin (red). DAPI was used to stain the cell nuclei (blue).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunocytochemistry analysis of LC3B in treated U373-MG cells. Samples were incubated in LC3B polyclonal antibody (Product # PA1-46286). Analysis using the HRP conjugate of anti-LC3B . The nuclei were stained with DAPI.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunocytochemistry analysis of LC3B in HeLa cells. Samples were incubated in LC3B polyclonal antibody (Product # PA1-46286). Antibody (red). Nuclei were counterstained with DAPI (blue).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunocytochemistry analysis of LC3B in treated U373-MG (human glioblastoma) cells. Samples were incubated in LC3B polyclonal antibody (Product # PA1-46286).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical analysis of LC3B in treated U87-MG cultured & subcutaneous tumors. Samples were incubated in LC3B polyclonal antibody (Product # PA1-46286).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical analysis of LC3B in gliobastoma multiform tissue. Samples were incubated in LC3B polyclonal antibody (Product # PA1-46286).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: FACS staining of NTERA-2 cells using Product # PA1-46286 at a 1:200 dilution detected using Dylight-488 conjugated goat anti-rabbit IgG secondary

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 7 Loperamide, pimozide, and STF-62247 enhance the autophagic flux in MZ-54 cells. a MZ-54 cells were treated with 20 uM IM/100 uM TIC, 17.5 uM loperamide, 15 uM pimozide, and 40 uM STF-62247 for 8, 2, 4 and 3 h, respectively. BafA1 was added 4 h before cell harvesting at a final concentration of 40 nM. Western blotting was performed with the indicated antibodies and vinculin as loading control. For quantification, LC3-II band intensities were normalized to vinculin band intensities. b Schematic representation of the GFP-LC3B-RFP-LC3BDeltaG autophagy flux sensor. Upon expression, the GFP-LC3B-RFP-LC3BDeltaG fusion protein is cleaved by the ATG4 proteases after which GFP-LC3B becomes conjugated to PE and localizes to autophagosomes which eventually fuse with lysosomes, inducing degradation of GFP-LC3B. RFP-LC3BDeltaG remains in the cytosol, where it serves as internal control. Scheme adapted from Kaizuka et al. 48 c Stable GFP-LC3B-RFP-LC3BDeltaG-expressing MZ-54 WT or ATG7 KO cells were treated as indicated in a for 16 h followed by Western blotting with vinculin as loading control. d , e Stable GFP-LC3B-RFP-LC3BDeltaG-expressing MZ-54 WT ( d ) or ATG7 KO cells ( e ) were treated with 20 uM IM/100 uM TIC, 15 uM loperamide, 15 uM pimozide or 40 uM STF-62247 for the indicated time points followed by flow cytometry. BafA1 was added 4 h before the measurement at a final concentration of 40 nM. Mean and SEM of three independent experiments performed in triplicate

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 3 Representative western blots of LC3 and SQSTM1/p62 proteins in the anterior tibialis muscle of juvenile mice sacrificed at post-natal day (PND)136 showing the inter-individual and inter-group variation of protein expression: ( A ) Male data n = 5/group; ( B ) Female data n = 5/group. Standardized protein levels were analysed by the Kruskal-Wallis test. Mann-Whitney was used for multiple comparisons; * p < 0.05.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 4 Representative Western blot of LC3 and SQSTM1/p62 proteins in the anterior tibialis muscle of adult mice sacrificed at PND136 showing the inter-individual and inter-group variation of protein expression: ( A ) Male data n = 5/group; ( B ) Female data n = 5/group. Standardized protein levels were analysed by Kruskal-Wallis test. Mann-Whitney was used for multiple comparisons; * p < 0.05.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 7 4PYR induced autophagy in HepG3 cells. HepG3 cells were continuously dosed with 100 uM of 4PYR for 48, 72, and 96 h. ( A ) Increased protein expression levels of the autophagy marker LC3BII, Beclin-1, and decrease in phospho-mTOR were assessed using immunoblot in 4PYR treated HepG3 cells compared to vehicle-treated control cells. alpha-tubulin was used as a loading control for immunoblotting. The graph shows quantified protein expression levels relative to controls for ( B ) LC3B-II, ( C ) Beclin-1 and ( D ) phosphor-mTOR in HepG3 cells. Results are expressed as relative abundance (a.u.) +- SEM of two biological replicates. Statistical significance: * p < 0.05.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 3 Evidence of placental autophagy induction after suppressing the CXCL12-CXCR4 network. Protein abundance of cellular autophagy marker LC3B-II and representative immunoblots in ovine fetal cotyledon (COT) placenta (A) and maternal caruncle (CAR) placenta (B) following intrauterine saline (CON) or AMD3100 (AMD) infusion. Data are presented as the mean +- SEM, and significance is denoted with an asterisk when p < 0.05 (*).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2 STF-62247 and pimozide lead to a strong accumulation of endogenous LC3B-II protein in MEFs. ( A , B ) Atg5 +/+ and Atg5 -/- MEFs were treated with 20 uM STF-62247, 15 uM pimozide or 7.5 uM ABT-737/10 uM etoposide for 24 hours ( A ). Atg7 +/+ and Atg7 -/- MEFs were treated with 40 uM STF-62247, 10 uM pimozide or 7.5 uM ABT-737/10 uM etoposide for 24 hours ( B ). Formation of LC3B puncta was imaged using anti-LC3B immunofluorescence staining. Representative images over 25 sites per sample are shown. ( C , D ) Quantification of mean LC3B puncta per cell upon STF-62247, pimozide or ABT-737/etoposide treatment of Atg5 +/+ and Atg5 -/- ( C ) or Atg7 +/+ and Atg7 -/- ( D ) MEFs. Mean and SEM of three independent experiments performed for 25 sites per sample are shown. Significances after drug treatment of Atg5 +/+ , Atg5 -/- and Atg7 +/+ , Atg7 -/- cells are calculated versus untreated cells of the corresponding cell line. Scale bar = 30 uM. **p < 0.01, ***p < 0.001. ut = untreated, STF = STF-62247, PIMO = pimozide, eto = etoposide.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 3 STF-62247 and pimozide lead to enhanced autophagic flux in MEFs. ( A , C ) Atg5 +/+ MEFs were treated for 4 hours with 20 uM STF-62247 ( A ) or 15 uM pimozide ( C ) in the absence or presence of 40 nM BafA1. Protein levels were detected by Western blotting with vinculin as loading control. ( B , D ) Atg7 +/+ MEFs were treated for 16 hours with 40 uM STF-62247 ( B ) or for 4 hours with 10 uM pimozide ( D ) in the absence or presence of 40 nM BafA1. Protein levels were detected by Western blotting with vinculin as loading control. For quantification, LC3B-II protein levels were normalized to vinculin protein levels and expressed as fold changes compared to the untreated sample. ( E , F ) Atg5 +/+ and Atg7 +/+ MEFs were transfected with mRFP-EGFP-LC3B followed by treatment with 20 uM STF-62247 or 15 uM pimozide and 40 uM STF-62247 or 10 uM pimozide in the absence or presence of 40 nM BafA1, respectively, for 8 hours ( Atg5 +/+ ) or for 12 hours ( Atg7 +/+ ). Images were acquired by confocal microscopy. Arrows highlight mRFP + /EGFP - puncta. ( G , H ) The numbers of mRFP + /EGFP - and mRFP + /EGFP + dots per cell were quantified after treatment of Atg5 +/+ ( G ) and Atg7 +/+ ( H ) MEFs with 20 uM STF-62247 or 15 uM pimozide and 40 uM STF-62247 or 10 uM pimozide, respectively, in the absence and presence of 40 nM BafA1. Mean and SEM of three independent experiments are shown. 21-51 cells were quantified per sample. Red stars indicate significances of mRFP + /EGFP - dots

PA1-46286

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