ATG2A (#15011), ATG5 (#2630), ATG7 (#2631), BECN1 (#3738), BIRC2 (D5G9), CASP8 (1C12), FADD (#2782), FAS (C18C12), FASLG (#4273), HSP90 (C45G5); LAMP1 (C54H11), MLKL (D2I6N), PARP1 (46D11), RIPK1 (D94C12), RIPK3 (D4G2A), ULK1 (D9D7), and XIAP (3B6) antibodies were obtained from Cell Signaling. SQSTM1 (#ab56416) and STX17 (#ab116113) antibodies were purchased from Abcam. β-actin (ACTB; AC-74) and ATG2B (#HPA019665) were purchased from Sigma. MAP1LC3B (#NB100-2220) was purchased from Novus Biologicals. Cell lysis, co-immunoprecipitation, subcellular fractionation, and western blotting were performed as previously described19 (link),41 (link). Relative densities of the target bands to the reference bands was calculated using ImageJ (NIH).
Map1lc3b
Manufactured by Novus Biologicals
Sourced in United States
MAP1LC3B is a protein that is involved in the process of autophagy, which is a cellular mechanism responsible for the degradation and recycling of damaged or unnecessary cellular components. This protein plays a crucial role in the formation of autophagosomes, which are the vesicles that engulf and transport cargo to the lysosome for degradation. MAP1LC3B is commonly used as a marker for monitoring autophagic activity in cells.
Automatically generated - may contain errors
Lab products found in correlation
Becn1, by Cell Signaling Technology (2 mentions)
Hoechst 33342, by Thermo Fisher Scientific (2 mentions)
Xf plasmamembrane permeabilizer xf pmp, by Agilent Technologies (2 mentions)
Nucleofector kit 5, by Lonza (2 mentions)
Paraformaldehyde (pfa), by Electron Microscopy Sciences (2 mentions)
Normal goat serum (ngs), by Merck Group (2 mentions)
β actin, by Merck Group (2 mentions)
Vps28, by Santa Cruz Biotechnology (2 mentions)
Membrane impermeable halotag ligand mil, by Promega (2 mentions)
Membrane permeable halotag ligand mpl, by Promega (2 mentions)
03 gp62 c, by Abcam (2 mentions)
Tsg101, by Abcam (2 mentions)
Bafilomycin a1, by LC Laboratories (2 mentions)
Xiap 3b6, by Cell Signaling Technology (1 mentions)
Atg2b, by Merck Group (1 mentions)
Atg2a, by Cell Signaling Technology (1 mentions)
Sqstm1 ab56416, by Abcam (1 mentions)
Pecam 1, by BD (1 mentions)
Hif 1a, by Novus Biologicals (1 mentions)
Chemmate dako envision kit, by Agilent Technologies (1 mentions)
7 protocols using map1lc3b
1
Autophagy-related Protein Detection
2
Comprehensive Antibody Validation Protocol
3
Immunohistochemical Analysis of Tumor Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Sections of tumor tissue blocks were cut onto adhesive-coated glass slides (Instrumedics, Hackensack) at a thickness of 3 μm. The slides were dewaxed in xylene and rehydrated through a graded alcohol series. Then, slides were pressure-cooked in 10 mM citrate buffer (pH 6) for 7 minutes for antigen retrieval and washed using TBS with 0.1% Tween 80 for 5 minutes. Endogenous peroxidase activity was blocked by 3% H2O2 treatment. After washing, the slides were incubated with primary antibodies against KIT (Epitomics), MAP1LC3B, HIF1A (Novus Biologicals), and PECAM1 (BD Biosciences) at RT for 1 hour. Primary antibodies were detected following the user's manual of the ChemMate DAKO EnVision kit (DAKO). The slides were incubated with the secondary antibody for 30 minutes and developed with 3,3-diaminobenzidine for 5 minutes. Slides were then counterstained with hematoxylin. Incubation without the primary antibody was used as a negative control.
4
Comprehensive Antibody Validation Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The following antibodies were used for immunoblotting (IB) and immunofluorescence (IF): β-ACTIN (IB, Sigma-Aldrich, A5441, 1:10,000); GFP (IB, Cell Signaling, 2956, 1:1,000); HA (IB, IF, BioLegend, 901513, 1:2,000 (IB), 1:1,000 (IF)); MAP1LC3B (IB, Novus, NB100–2220, 1:3,000; IF, Cell Signaling, 3868, 1:200); p62 (IB; American Research Products, 03-GP62-C, 1:4,000); TSG101 (IB, Abcam, ab83, 1:1,000); VPS28 (IB, Santa Cruz Biotechnology, sc-166537, 1:100). ON-TARGETplus SMART Pool Non-targeting (D-001810–10) and CHMP2A (L-020247–01) siRNAs were obatined from GE Healthcare Dharmacon. pCDH1-CMV-HA-VPS28(WT)-SV40-hygro and pCDH1-CMV-HA-VPS28(TM[K54D, K58D, D59A])-SV40-hygro were generated using Gibson Assembly. All other reagents were obtained from the following sources: Bafilomycin A1 (LC Laboratories, B-1080); Hoechst 33342 (Invitrogen, NucBlue, R37605); Membrane-impermeable HaloTag Ligand (MIL) (Promega, Alexa Fluor 488-conjugated, G1001); Membrane-permeable HaloTag Ligand (MPL) (Promega, tetramethylrhodamine-conjugated, G8251); normal goat serum (Sigma-Aldrich, G9023); Nucleofector Kit V (Lonza, VCA-1003); paraformaldehyde (Electron Microscopy Sciences, 15710); XF Plasma Membrane Permeabilizer (XF-PMP) (Seahorse Bioscience, 102504–100).
5
Immunocytochemical Analyses of Autophagy
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Immunocytochemistry was performed as previously described (Bekbulat et al, 2020 (link)). Briefly, cells were grown on glass coverslips, fixated with 4% PFA, and permeabilized with 90% (v/v) methanol. Unspecific binding sites were blocked with 3% BSA in PBS followed by incubation with primary antibody, fluorophore‐conjugated secondary antibody, and DAPI. For sphingomyelin analyses, cells were treated with 500 nM BODIPY FL C5‐shingomyelin (Fisher, 11510306) in serum‐free EBSS for 1 h and thereafter were incubated for 1 h with bafilomycin A1 in EBSS. Cells were imaged using the laser scanning microscope LSM710 (Zeiss). Primary antibody: MAP1LC3B (Nanotools, 0260‐100/LC3‐2G6); MAP1LC3B (Novus, NB100‐2220); SQSTM1 (Progen, GP62‐C); LAMP2 (Abcam, ab25631); TGN46 (BioRad, AHP500); HA (Sigma, H6908). Secondary antibody: Cy3 anti‐mouse (ImmunoResearch, 715‐165‐151), Cy3 anti‐rabbit (ImmunoResearch, 711‐165‐152), Cy3 anti‐sheep (ImmunoResearch, 713‐165‐147), Cy5 anti‐mouse (ImmunoResearch, 715‐175‐150), Cy5 anti‐rabbit (ImmunoResearch, 711‐175‐152).
6
Antibody Expression Analysis in Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The following antibodies were used: CCR5 (Cat# ab65850, RRID:AB_1140936), CXCR4 (Cat# ab124824, RRID:AB_10975635), and SQSTM1 (Cat# ab56416, RRID:AB_945626) from Abcam, CD4 (Cat# 300501, RRID:AB_314069) from BioLegend, ATG5 (Cat# 2630, RRID:AB_2062340), ATG7 (Cat# 2631, RRID:AB_2227783), BIRC2 (Cat# 7065, RRID:AB_10890862), CASP8 (Cat# 9746, RRID:AB_2275120), PARP1 (Cat# 9532, RRID:AB_659884), RIPK1 (Cat# 3493, RRID:AB_2305314), XIAP (Cat# 2045, RRID:AB_2214866) from Cell Signaling Technologies, MAP1LC3B (Cat# NB100-2220) from Novus Biologicals, and ACTB (Cat# A2228, RRID:AB_476697) from Sigma. Whole cell lysates and TNP membranes were prepared, resolved, and proteins detected as previously described 10 (link), 52 (link). Relative densities of the target bands were compared to the reference (ACTB) and were calculated using Fiji from the Max Planck Institute of Molecular Cell Biology and Genetics (Fiji, RRID:SCR_002285) 53 (link).
7
Antibody and Chemical Treatments for Autophagy Study
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The following antibodies were used in this study: acetylated lysine (Cell Signaling; Danvers, MA, USA; 9441), BECN1 (Cell Signaling, 3738), BNIP3 (Santa Cruz Biotechnology, Dallas, TX, USA; sc-56167), GAPDH (Santa Cruz Biotechnology; sc-137179), GAC (GeneTex, Irvine, CA, USA; GTX131263), HIF-1α (Cell Signaling; 14179), MAP1LC3B (Novus Biologicals, Centennial, CO, USA; NB600-1384), PRKN (Santa Cruz Biotechnology, sc-32282), SLC20A1/2 (PiT-1/2) (Santa Cruz Biotechnology, sc-101298), SLC25A3 (Santa Cruz Biotechnology, sc-376742), ULK1 (Abcam, Cambridge, UK; ab128859), VDAC1 (Santa Cruz Biotechnology, sc-390996), pULK1 (Abcam; ab156920), peroxidase-conjugated AffiniPure goat anti-rabbit IgG (H + L) (Jackson ImmunoResearch, West Grove, PA, USA; 111-035-003), and peroxidase-conjugated AffiniPure goat anti-mouse IgG (H + L) (Jackson ImmunoResearch; 115-035-062).
Lanthanum acetate (Merck; 306339) was dissolved in distilled water, filtered and then added to a final concentration of 2 mM every 24 h and 2 h before harvesting the cells.
Cobalt (III) chloride hexahydrate (Merck; C8661) was dissolved in distilled sterile water and added to a final concentration of 200 µM for 24 h or 48 h before harvesting the cells.
Bafilomycin A1 (Merck, B1793) was dissolved in dimethyl sulfoxide (DMSO) and added to a final concentration of 100 nM for 2 h.
Lanthanum acetate (Merck; 306339) was dissolved in distilled water, filtered and then added to a final concentration of 2 mM every 24 h and 2 h before harvesting the cells.
Cobalt (III) chloride hexahydrate (Merck; C8661) was dissolved in distilled sterile water and added to a final concentration of 200 µM for 24 h or 48 h before harvesting the cells.
Bafilomycin A1 (Merck, B1793) was dissolved in dimethyl sulfoxide (DMSO) and added to a final concentration of 100 nM for 2 h.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!