Baf a1

Manufactured by MedChemExpress

Sourced in United States

Baf-A1 is a laboratory reagent that functions as a potent and specific inhibitor of vacuolar-type H+-ATPase (V-ATPase). It is commonly used in cell biology research to investigate the role of V-ATPase in various cellular processes.

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Lab products found in correlation

Rapamycin, by MedChemExpress (3 mentions) Z vad fmk, by MedChemExpress (2 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (2 mentions) Sq22536, by Selleck Chemicals (1 mentions) Anaeropack system, by Mitsubishi (1 mentions) Hemin, by Merck Group (1 mentions) Dulbecco modified eagle medium (dmem), by Solarbio (1 mentions) Foetal bovine serum (fbs), by Yeasen (1 mentions) Sp600125, by MedChemExpress (1 mentions) Hrp conjugated goat anti rabbit igg, by ZSGB-BIO (1 mentions) Sb203580, by MedChemExpress (1 mentions) Chloroquine, by MedChemExpress (1 mentions) Hydroxychloroquine, by MedChemExpress (1 mentions) Glomax 96 microplate luminometer, by Promega (1 mentions) Rapamycin, by Selleck Chemicals (1 mentions) Dc661, by MedChemExpress (1 mentions) Celltiter glo luminescent cell viability assay, by Promega (1 mentions) Transit lt1 transfection reagent, by Mirus Bio (1 mentions) Phenanthroline, by Merck Group (1 mentions) Clozapine, by Merck Group (1 mentions)

Close spelling variants of this product

Bafilomycin A1 (Baf A1) (18 citations) Bafilomycin A1 (BAF), (2 citations)

13 protocols using baf a1

Endothelial Cell Hypoxia and Glucose Modulation

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We created HG/Hypo conditions by adding 33.3 mM glucose to the endothelial cell medium with 1% FBS, and the Anaeropack system (Cat# D-07, Mitsubishi Gas Chemical Co, Japan) was used to create a hypoxia condition. Next, HUVECs in a complete endothelial cell medium with 5.5 mM glucose without the Anaeropack system were cultured as the control group (normal glucose, NG). After HUVECs were cultured for 2 days under NG and HG/Hypo conditions, MoS₂ NDs (50 µg/mL) or PBS were added to the culture medium for 12 h for subsequent experiments. HUVECs were incubated with Baf A1 (1 nM) (Cat# HY-100558, MedChemExpress, USA), 3-MA (Cat#HY-19132, MedChemExpress, USA) and SQ22536 (250 µM) (Cat# S8283, Selleck, USA) to inhibit autophagy and cAMP level, respectively.

Wang Y., Li F., Mao L., Liu Y., Chen S., Liu J., Huang K., Chen Q., Wu J., Lu L., Zheng Y., Shen W., Ying T., Dai Y, & Shen Y. (2024). Promoting collateral formation in type 2 diabetes mellitus using ultra-small nanodots with autophagy activation and ROS scavenging. Journal of Nanobiotechnology, 22, 85.

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Cellular Responses to Hemin, Baf-A1, and ZnPP

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BV‐2 and HT22 were obtained from the China Center for Type Culture Collection and cultured in DMEM (11995; Solarbio Co., Ltd.) containing 10% foetal bovine serum (40130ES76; Yeasen Co., Ltd.), supplemented in acceptable condition at 37°C, 5% CO₂. Cells were seeded at 5 × 10⁴ cells/mL. When the cells were confluent to 70%, hemin (H9039; Sigma‐Aldrich Co., Ltd.), Baf‐A1 (HY‐100558; MedChemExpress Co., Ltd.) and ZnPP (HY‐101193; MedChemExpress Co., Ltd.) were used in this experiment.

Wang R., Liang Z., Xue X., Mei H., Ji L., Wang B., Chen W., Gao C., Yuan S., Wu T., Qi H., Hu S., Yi L., Song Y., Liao R, & Chen B. (2023). Microglial FoxO3a deficiency ameliorates ferroptosis‐induced brain injury of intracerebral haemorrhage via regulating autophagy and heme oxygenase‐1. Journal of Cellular and Molecular Medicine, 28(1), e18007.

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Molecular Signaling Pathway Modulation

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P38 MAPK inhibitor SB203580, ERK inhibitor U1026, JNK inhibitor SP600125, Baf-A1 and Rapamycin were obtained from MedChemExpress (Shanghai, China). The above reagents were dissolved in dimethyl sulfoxide (DMSO) in the experiments. Rabbit anti-p-p53 (80195-1-RR), anti-p53 (10442-1-AP) antibody was obtained from Proteintech (Wuhan, China). The other antibodies used in the experiments were purchased from Cell Signaling Technology (Maryland, USA). The HPR-conjugated goat anti-mouse IgG and HRP-conjugated goat anti-rabbit IgG antibodies were purchased from ZSGB-BIO (China). siRNAs for negative control and JNK were purchased from Ribobio (Guangzhou, China).

Cui X., Chen F., Zhao J., Li D., Hu M., Chen X., Zhang Y, & Han L. (2023). Involvement of JNK signaling in Aspergillus fumigatus-induced inflammatory factors release in bronchial epithelial cells. Scientific Reports, 13, 1293.

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Cytotoxicity Assay of Small Molecule Inhibitors

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Most cells were obtained from American Type Culture Collection (ATCC) and were cultured in ATCCrecommended media supplemented with 10% fetal bovine serum and 2% antibiotics (Penicillin-Streptomycin). MDA-MB-231 CRISPRi cells were a gift from Hani Goodarzi at UCSF. MEFs expressing human KRAS/BRAF were from the Frederick National Laboratory for Cancer Research, The National Cancer Institute. Baf A1, Chloroquine, HydroxyChloroquine, and DC661 were from Medchemexpress LLC. Rapamycin was from Selleckchem and Torin was from Fisher. DNA transfection into cells was performed with TransIT®-LT1 Transfection Reagent (Mirus Bio) by following the manufacturer's recommendations.
Logarithmically growing cells were plated in antibiotic-free medium supplemented with 2% fetal bovine serum at a density of 5,000 cells per well. The next day, cells were treated in triplicates with increasing doses of in-house small molecule inhibitor drugs and a DMSO vehicle control for 3 days and subsequently assessed for cell viability by measurement of ATP with CellTiter-Glo Luminescent Cell Viability Assay (Promega). Signal intensity was read on a Glomax TM 96 Microplate Luminometer (Promega) and percent cell survival was calculated based on the reading of vehicle control cells set as 100%. Each compound was tested a minimum of 2 times with n = 3.

Tolani B., Celli A., Yao Y., Tan Y.Z., Fetter R.D., Liem C.R., Smith A.J.d., Vasanthakumar T., Bisignano P., Seiple I.B., Rubinstein J.L., Jost M., & Weissman J.S. (2020). Selective Targeting of Ras Mutant Cancers via a New Small Molecule.

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Multiparametric Analysis of Cellular Responses

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All reagents were dissolved and stored in DMSO or water. Baf-A1, Rapamycin, Torin-1, zVADfmk was obtained from Med Chem Express. DQ-BSA-red, FluoZin-3 AM, LysoTracker Red DND-99, BAPTA-am, EDTA, PI, and H2DCFDA were from Life Technologies. Clozapine, FTY720, NS8593, TTM, DIP, Annexin V/7-AAD, CQ, 3-MA, TPEN, 1,10 Phenanthroline were purchased from Sigma. Trypan blue was purchased from VWR. Naltriben was purchased from Santa Cruz Biotechnology.
Statistical Analysis. Data are presented as the means ± standard errors of the mean (SEMs) from at least three independent experiments. Statistical signi cance of differences was evaluated using ANOVA followed by Tukey's test. P values < 0.05 were considered statistically signi cant.

Xing Y., Wei X., Wang M.M., Liu Y., Sui Z., Wang X., Zhang Y., Fei Y.H., Jiang Y., Lu C., Zhang P., Chen R., Liu N., Wu M., Ding L., Wang Y., Guo F., Cao J.L., Qi J, & Wang W. (2022). Stimulating TRPM7 suppresses cancer cell proliferation and metastasis by inhibiting autophagy. Cancer letters, 525.

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Autophagy and Cell Death Pathway Assay

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Reagents utilized in this study included: 3-Methyladenine (3-MA; Millipore Sigma, M9281), bafilomycin A1 (Baf-A1; MedChem Express, HY-100558), Chloroquine (CQ; MedChem Express, HY-17589A), Z-VAD-FMK (Z-VAD; MedChem Express, HY-16658B), Necrostatin-1 (Nec-1; MedChem Express, HY-15760), Ferrostatin-1 (Fer-1; MedChem Express, HY-100579), Liproxstatin-1 (Lip-1; MedChem Express, HY-12726) and R162(MedChem Express, HY-103096). Antibodies used in this study were: LC3B (Milliopore Sigma, L7543), SQSTM1/p62 (Milliopore Sigma, P0067), GAPDH (Diagbio, db106), HuR/ELAVL1 (ABclonal Technology, A19622), GLUD1 (Santa Cruz Biotechnology, sc-515542), AGO2 (ABclonal Technology, A19709) and Flag (MultiSciences, 70-AB002-100).

Zhu T., Cen Y., Chen Z., Zhang Y., Zhao L., Wang J., Lu W., Xie X, & Wang X. (2022). Oncogenic circTICRR suppresses autophagy via binding to HuR protein and stabilizing GLUD1 mRNA in cervical cancer. Cell Death & Disease, 13(5), 479.

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Cloning and Characterization of Zebrafish Immune Genes

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Zebrafish gene mylipb (ENSDARG00000055118) and its truncated mutants were PCR amplified from AB zebrafish cDNA using PCR. Amplified genes were subcloned into pCMV-Myc (Clontech), pCMV-HA (Clontech), or pCMV-Flag (Clontech) vectors. These plasmids, including Myc-rig-i, Myc-mda5, Myc-tbk1, Myc-irf3, were constructed using the pCMV-Myc vector as described previously. All constructs were verified by DNA sequencing.
VigoFect (Vigorous Biotechnology, Beijing, China) and FishTrans (MST, FT2020, Wuhan, China) were used for cell transfection. poly I:C (InvivoGen, San Diego, CA) was transfected with Lipofectamine 3000 (Thermo Fisher Scientific). Cycloheximide (CHX), MG132, Baf-A1 and 3-MA were purchased from MedChem Express. NH₄Cl was purchased from Sigma-Aldrich. The antibodies used were as follows: anti-Flag antibody (F1804; Sigma-Aldrich), anti-Myc antibody (9E10; Santa Cruz Biotechnology), anti-HA antibody (MMS-101R; Covance), anti-LC3 antibody (ab48394; Abcam), anti-irf3 (A11921, Abclonal), anti-phosphor-IRF3 (4947, Cell Signaling Technology). anti-ATG5 (A0203, ABclonal), anti-BECN1 (A17028, ABclonal), anti-α-tubulin (62204, Thermo Fisher Scientific), anti-GAPDH antibody (AC033; ABclonal) and anti-β-actin antibody (AC026; ABclonal).

Li Z., Li J., Li Z., Song Y., Wang Y., Wang C., Yuan L., Xiao W, & Wang J. (2024). Zebrafish mylipb attenuates antiviral innate immunity through two synergistic mechanisms targeting transcription factor irf3. PLOS Pathogens, 20(5), e1012227.

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Cell Line Culture and Compound Screening

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Aspc-1, HepG2, Panc02, and H22 cell lines were obtained from the KeyGEN Biotechnology Company (China). HT1080 and SW480 were obtained from the FuHeng BioLogy Company (China). HT1080 cancer cells were cultured in Eagle’s Minimum Essential Medium supplemented with 10% fetal bovine serum (FBS), glutamine (2 mM), penicillin (100 U/ml), and streptomycin (0.1 mg/ml). SW480, Aspc-1, HepG2, Panc02, and H22 were cultured in high Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% FBS, L-glutamine (4 mM), and penicillin (100 U/ml) and streptomycin (0.1 mg/ml). All cell lines were maintained in a humidified atmosphere containing 5% CO₂ at 37 °C and tested for mycoplasma prior to the commencement of experiments. Unless otherwise indicated, cell culture medium was changed every 3 days, and cells were passaged using 0.05% trypsin/EDTA. Erastin (#HY-15763), sorafenib (#HY-10201), sulfasalazine (#HY-14655), DON (#HY-108357), RSL3 (#HY-100218A), L-Buthionine-(S,R)-sulfoximine (BSO, #HY-106376A), ferrostatin-1 (#HY-100579), Z-VADFMK (#HY-16658), AICAR (#HY-13417), BafA1 (#HY-100558), N-Acetylcysteine (#HY-B0215), and Necrosulfonamide (#HY-100573) were purchased from MedChemExpress (USA). Compound C (#ab120843) was purchased from Abcam. Deferoxamine mesylate (#D9533) was purchased from Sigma-Aldrich.

Wang K., Zhang Z., Tsai H.I., Liu Y., Gao J., Wang M., Song L., Cao X., Xu Z., Chen H., Gong A., Wang D., Cheng F, & Zhu H. (2020). Branched-chain amino acid aminotransferase 2 regulates ferroptotic cell death in cancer cells. Cell Death and Differentiation, 28(4), 1222-1236.

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Autophagy Regulation by EZH2 Inhibition

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We purchased anti-microtubule-associated protein LC3, P62, CD63, and BECN1 (beclin1, an autophagosome initiator) antibodies from R&D Systems (Minneapolis, MI, United States). Cell Signaling Technology (Danvers, MA, United States) provided the following antibodies: anti-EZH2 (#5246), anti-mTOR (#2983), anti-p-mTOR (#5536), anti-S6K1 (#2708), anti-p-S6K1 (#9204), anti-TSG101 (#28405), and anti-Ki67 (#9449). GSK126 (EZH2inhibitor) (10 μM), PQR620 (50 nM) and bafilomycin A1 (Baf A1, an autophagosome-lysosome fusion inhibitor) (100 nM) were purchased from MedChem Express (Monmouth Junction, NJ, United States). EZH2 plasmids were purchased from Shanghai Genechem Co., Ltd.

Chu Y., Chen W., Peng W., Liu Y., Xu L., Zuo J., Zhou J., Zhang Y., Zhang N., Li J., Liu L., Yao K., Gao G., Wang X., Han R., Liu C., Li Y., Zhou H., Huang Y, & Ye Y. (2020). Amnion-Derived Mesenchymal Stem Cell Exosomes-Mediated Autophagy Promotes the Survival of Trophoblasts Under Hypoxia Through mTOR Pathway by the Downregulation of EZH2. Frontiers in Cell and Developmental Biology, 8, 545852.

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Regulation of TBK1 by A137R

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HEK293T cells were cotransfected p3×Flag-TBK1 with pMyc-A137R or pCMV-Myc for 24 h. Then the cells were treated with 20 μM proteasome inhibitor MG132 (catalog no. M8699-1MG; Sigma-Aldrich), 10 mM autophagosome inhibitor 3-MA (catalog no. HY-19312; Medchemexpress), or 100 nM BafA1 (catalog no. HY-100558; Medchemexpress) for 8 h, and the lysates were examined by Western blotting analysis.

Sun M., Yu S., Ge H., Wang T., Li Y., Zhou P., Pan L., Han Y., Yang Y., Sun Y., Li S., Li L.F, & Qiu H.J. (2022). The A137R Protein of African Swine Fever Virus Inhibits Type I Interferon Production via the Autophagy-Mediated Lysosomal Degradation of TBK1. Journal of Virology, 96(9), e01957-21.

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