Autophinib

Manufactured by Selleck Chemicals

Sourced in China, United States

Autophinib is a versatile laboratory equipment designed for automated sample processing. It features a high-throughput and precise liquid handling system for preparing and dispensing samples, enabling efficient and reproducible experimental procedures.

Automatically generated - may contain errors

Lab products found in correlation

5 protocols using autophinib

Transwell Assay for CD19 CAR-T Cell Migration

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The human CD19 CAR‐T cells (2 × 10⁶ cells /ml, 200 μL per well) in the X‐VIVO 15 medium were added to the top chambers of the 24‐well Transwell plates (80 μm pores; Corning, Lowell, MA, USA). The Nalm6 (10⁶/mL) and Raji (10⁶/mL) cells were cultured in RPMI 1640 medium (with 10% FBS) with autophinib (2 μmol/L, Selleck, Shanghai, China) and rapamycin (0.1 nmol/L, Selleck, Shanghai, China) for 72 hours. The Nalm6 (10⁶/mL) and Raji (10⁶/mL) cells with sgControl and RB1CC1 knockout were cultured in RPMI 1640 medium (with 10% FBS) for 72 hours. The culture supernatants were collected and added to the bottom chambers (600 μL per chamber). After 4 hours, the migrated CD19 CAR‐T cells were counted, and migration was quantified by flow cytometry.

Tang L., Zhang H., Zhou F., Wei Q., Du M., Wu J., Li C., Luo W., Zhou J., Wang X., Chen Z., Zhang Y., Huang Z., Wu Z., Wen Y., Jiang H., Liao D., Kou H., Xiong W., Mei H, & Hu Y. (2024). Targeting autophagy overcomes cancer‐intrinsic resistance to CAR‐T immunotherapy in B‐cell malignancies. Cancer Communications, 44(3), 408-432.

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RNA-seq Analysis of RB1CC1 Knockout Nalm6 Cells

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After the Nalm6 cells were treated with vehicle and autophinib (Selleck, S8596, 2 μmol/L) for 72 hours, the sgControl and RB1CC1 knockout (RB1CC1^KO) Nalm6 cells were collected for RNA sequencing. RNA extraction, library construction and sequencing were performed by the Beijing Novogene Company (Beijing, China). For bulk RNA sequencing analysis of bone marrow samples from patients treated with CD19 CAR‐T cell therapy, we analyzed the data set from GSE153670. The differentially‐expressed genes were defined as genes with a P value <0.05 and a log₂ fold change (log₂FC) of >1. The EdgeR package was used for data normalization and gene variance analysis. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis and gene set enrichment analysis (GSEA) [27 (link)] were performed. Receiver operating characteristic (ROC) analysis was conducted to identify a threshold gene set score with high positive and negative predictive values of clinical outcomes in the sample set. Visualization was performed using the R package ggplot2.

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Modulating Autophagy in Skin Regeneration

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To investigate whether and how autophagy influences skin regeneration during tissue expansion, we used rapa (S1039, Selleckchem, USA) and autophinib (S8596, Selleckchem, USA) to modulate autophagic activities in the expanded skin. autophinib inhibits autophagy, whereas rapa promotes autophagy [25 (link),27 (link)]. The animal model was established as described above. An area of 1 × 1 cm² of the scalp was marked on the tissue expander for tracing the expanded skin. Rapa, autophinib or rapa + autophinib were dissolved in DMSO (D8418, Sigma-Aldrich, USA). DMSO (vehicle), rapa, autophinib and rapa + autophinib, were individually mixed with pluronic lecithin organogel [16 (link),28 (link)] for topical application on the marked expanded skin at the time of each injection. The rats were divided into the following four groups (n = 6): control group (DMSO treatment), rapa group (1.6 μM), rapa + autophinib group (1.6 μM + 4 mM) and autophinib group (4 mM). The marked expanded skin specimens were harvested 48 h after the last expansion (day 28).

Du J., Liu W., Song Y., Zhang Y., Dong C., Xiong S., Huang Z., Wang T., Ding J., He Q., Yu Z, & Ma X. (2024). Activating autophagy promotes skin regeneration induced by mechanical stretch during tissue expansion. Burns & Trauma, 12, tkad057.

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AML Cell Line Profiling Protocol

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HL60, MOLM-14, MV4-11, OCI-AML2, OCI-AML3, U937, K562, THP1, and KASUMI AML cell lines were used (descriptions are provided in Supplemental Table 2). All AML cell lines were certified using their microsatellite identity and tested for mycoplasma contamination. Cells were cultured in RPMI (Gibco61870, Life Technologies® Saint Aubin, France) and supplemented with 10% fetal bovine serum (FBS) and 4 mM glutamine. VPS34 IN-1 was sourced from MRCC-PU Reagents (Dundee, Scotland). DMSO, chloroquine and doxycycline were obtained from Sigma-Aldrich (St. Louis, MO). Autophinib, PIK-III, Ferrostatine-1, Q-VAD-OPH, and necrostatine-1 were purchased from Selleckchem (Munich, Germany). Lysotracker deep red was obtained from Thermo Fischer Scientific (Asnières, France). l-Asparaginase was provided by Cochin hospital pharmacy Department.

Meunier G., Birsen R., Cazelles C., Belhadj M., Cantero-Aguilar L., Kosmider O., Fontenay M., Azar N., Mayeux P., Chapuis N., Tamburini J, & Bouscary D. (2020). Antileukemic activity of the VPS34-IN1 inhibitor in acute myeloid leukemia. Oncogenesis, 9(10), 94.

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Preparation and Use of Experimental Cell Culture Drugs

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Drugs for cell culture experiments were obtained as follows: buparlisib (SelleckChem; S2247), pictilisib (SelleckChem; S1065), copanlisib (SelleckChem; S2802), BX795 (SelleckChem; S1274), BX912 (SelleckChem; S1275), GSK2334470 (Tocris; 4143), bafilomycin A (Sigma-Aldrich; SML1661), 3-methyladenine (SelleckChem; S2767), autophinib (SelleckChem; S8596), and VPS34-IN1 (SelleckChem; 7980).
bafilomycin A was used at a concentration of 160 nM for all experiments. All other drug doses and incubation times are indicated in the figure legends or directly on the figures. Drugs were all dissolved in DMSO (Sigma-Aldrich; D2650), except for copanlisib and 3-methyladenine. copanlisib, which is not DMSO-soluble, was initially solubilized in a 10% trifluoroacetic acid (Sigma-Aldrich; T6508)/90% water solution to a concentration of 2 mM and then diluted to appropriate working stock concentrations. 3-methyladenine solutions were prepared fresh each time. 3-methyladenine powder was dissolved directly into cell culture medium to produce a 50 mM stock. 3-methyladenine solutions were heated to 55°C for 5 min, vortexed until clear, diluted to the appropriate concentration, and used immediately.
For all drug treatments, the cells were maintained in complete growth medium, with serum, according to the recipes specified above.

Endicott S.J., Ziemba Z.J., Beckmann L.J., Boynton DN J.r, & Miller R.A. (2020). Inhibition of class I PI3K enhances chaperone-mediated autophagy. The Journal of Cell Biology, 219(12), e202001031.

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