Gtx100685

Manufactured by GeneTex

GTX100685 is an electrophoresis unit designed for the separation and analysis of macromolecules, such as proteins and nucleic acids, based on their size and charge. It provides a controlled environment for the migration of samples through a gel matrix under the influence of an electric field. The core function of this product is to facilitate the electrophoretic separation of biomolecules for various analytical and preparative applications.

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Lab products found in correlation

Gtx127375, by GeneTex (2 mentions) Ab52894, by Abcam (1 mentions) Hy 13259, by MedChemExpress (1 mentions) Hy 100558, by MedChemExpress (1 mentions) Hy 10940, by MedChemExpress (1 mentions) Hy b1370, by MedChemExpress (1 mentions) Gapdh 6c5, by Santa Cruz Biotechnology (1 mentions) Bafilomycin a1, by Merck Group (1 mentions) 3 methyladenine, by Merck Group (1 mentions) Chloroquine diphosphate salt, by Merck Group (1 mentions) Radioimmunoprecipitation assay (ripa), by Solarbio (1 mentions) Se134, by Solarbio (1 mentions) Bicinchoninic acid (bca), by Thermo Fisher Scientific (1 mentions) Ab207612, by Abcam (1 mentions) Ab211033, by Abcam (1 mentions) Ab110304, by Abcam (1 mentions) Ab68477, by Abcam (1 mentions) Ab92946, by Abcam (1 mentions) Quantity one software program, by Bio-Rad (1 mentions) Sc 81178, by Santa Cruz Biotechnology (1 mentions)

6 protocols using gtx100685

Synthesis and Evaluation of Curcumin Derivatives

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The following were the antibodies used: anti-mouse tubulin (Genetex, GTX27291), anti-rabbit EGFR (abcam, ab52894), anti-rabbit pEGFR (abcam, ab5644), anti-rabbit hsp70 (Genetex, GTX104126), anti-rabbit bag3 (Genetex, GTX102396), and anti-rabbit p62 (Genetex, GTX100685). The following were the reagents used: ver-155008 (Medchem Express, HY-10941), pif (Medchem Express, HY-10940), bafilomycin A1 (Medchem Express, HY-100558), hydroxychloroquine (Medchem Express, HY-B1370), ixazomib (Medchem Express, HY-10453), MG132 (Medchem Express, HY-13259), osi (Medchem Express, HY-15772), and gef (LC lab, G-4408). The synthesis of all curcumin derivatives has been reported in our earlier publication (26 (link)). The structures and synthetic procedures of MTH-3, 21b, 22a, 22b, 23a, 24b, 25, 33, 35a, 35d, 36, and 37 correspond to the synthetic methods employed for the synthesis of compounds 9a, 10a, 9b, 10b, 11, 14a, 14b, 15, 16a, 16d, 17, and 18, respectively, as depicted in Scheme 1A-C of the reference (26 (link)). Briefly, curcumin derivatives were synthesized from curcumin via a three-step sequence involving esterification of curcumin with 2,2,5-trimethyl-1,3-dioxane-5-carboxylic acid to afford an ester intermediate, which was in turn dialkylated with alkyl bromide and hydrolysis with HCl(aq).

Hong X., Hsieh M.T., Tseng T.Y., Lin H.Y., Chang H.C., Yau S.T., Cheng W.C., Ke B., Liao H.H., Wu C.Y., Liu A.A., Wu M.M., Huang K.Y., Yang P.C., Kuo S.C., Hung M.C, & Lee P.C. (2023). Diarylheptanoid 35d overcomes EGFR TKI resistance by inducing hsp70-mediated lysosomal degradation of EGFR in EGFR-mutant lung adenocarcinoma. The Journal of Biological Chemistry, 299(6), 104814.

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Ras Protein and Autophagy Regulation

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The antibodies against N-Ras (F155), K-Ras (F234), H-Ras (F235), Cdc42(P1) and GAPDH (6C5) were from Santa Cruz Biotechnology. Antibodies against LC3B and p62/SQSTM1 were from Novus Biologicals (NB100–2220) and GeneTex (GTX100685), respectively. Flunarizine dihydrochloride (FLN) used in vitro, cycloheximide (CHX), MG132, bafilomycin A1, chloroquine diphosphate salt, and 3-methyladenine were all from Sigma-Aldrich. FLN used in vivo was from Medchemexpress.

Zheng Z.Y., Li J., Li F., Zhu Y., Cui K., Wong S.T., Chang E.C, & Liao Y.H. (2018). Induction of N-Ras degradation by flunarizine-mediated autophagy. Scientific Reports, 8, 16932.

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Protein Expression Analysis in MDBK Cells

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MDBK cells were lysed by radio immunoprecipitation assay (RIPA, Solarbio) lysis buffer with phenylmethylsulfonyl fluoride (PMSF, MCE). The concentration of proteins was measured using a bicinchoninic acid (BCA, Thermo Fisher Scientific) protein assay kit. The proteins were separated by SDS polyacrylamide gel electrophoresis (SDS-PAGE) and were transferred to polyvinylidenedifluoride (PVDF) membranes. The membranes were blocked in a nonfat dry milk solution for 30 min. Then, the membranes were incubated overnight at 4 ℃ with Notch3 (affinity, 1:1000 dilution), anti-mTOR (Bioss, bs-1992R, 1:1000 dilution), anti-p-mTOR (Affinity, AF3308, 1:1000 dilution), anti-LC3A (Novus Biologicals, NB100-2331, 1:1000 dilution), anti-p62 (GeneTex, GTX100685, 1:1000 dilution) and anti-Beclin1 (CST, 3495 T, 1:1000 dilution) antibody primary antibodies in dilution buffer. After washing with TBS-T, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-rabbit (Solarbio, SE134, 1:3000 dilution) at room temperature for 30 min. The membranes were developed using the ECL Western blot system (Tanon-5200) according to the manufacturer’ s instruction.

Cui Y., Guo H., Zhang Q., Fang J., Xie Y., Chen S., Ma X., Gou L., Cui H., Geng Y., Ye G., Zhong Z., Ren Z., Wang Y., Deng J., Yu S., Cao S., Wang Z, & Zuo Z. (2022). The combination of high glucose and LPS induces autophagy in bovine kidney epithelial cells via the Notch3/mTOR signaling pathway. BMC Veterinary Research, 18, 307.

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Western Blot Analysis of Autophagy and Stress Markers

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Proteins from liver tissues were extracted using RIPA buffer (containing 1% phenylmethylsulfonyl fluoride and 1% phosphatase inhibitors). The BCA protein assay kit (BL521A, Biosharp) was used to determine the protein concentration. Proteins were separated by SDS-PAGE and transferred into polyvinylidene fluoride (PVDF) membranes. The PVDF membranes were blocked with 5% non-fat dried milk in tris-buffered saline (pH 7.4) containing 0.1% Tween20 and subsequently immunoblotted with antibodies directed against BECN1 (ab207612, Abcam), ATG5 (GTX113309, GeneTex), MAP1LC3B (GTX127375, GeneTex), KHDRBS1 (GTX100685, GeneTex), HSPA5 (#3183, CST), DDIT3 (#2895, CST), SIRT1 (ab110304, Abcam), SIRT2 (ab211033, Abcam), NFE2L2(ab92946, Abcam), HMOX1 (ab68477, Abcam), cleaved-CASPASE 3 (#9661, CST), and β-ACTIN (sc-81178, Santa Cruz). The bands were visualized via an enhanced chemiluminescence kit (WBKLS0100, Millipore) and the semi-quantitative analysis was conducted by the Quantity One software program (Bio-Rad).

Li J., Zha X., Kang Y., Zhang Z., Yan L., Song L., Wang C, & Yang J. (2022). Oxygen-carrying sequential preservation mitigates liver grafts ischemia-reperfusion injury. iScience, 26(1), 105858.

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Western Blot Analysis of Apoptosis and Autophagy

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The western blot analyses were performed as described previously (Kuo et al., 2016). The protein concentrations of cell or tissue lysates were measured using the Lowry assay. Then protein samples (30 μg) were separated using 7.5%, 10%, or 12.5% SDS‐PAGE (according to the MW of the proteins of interest) and then electroblotted onto a nitrocellulose membrane. The membranes were blocked with 5% non‐fat dry milk in Tris‐buffered saline plus Tween, immunoblotted with specific primary antibodies against Bcl‐2 (1:500, sc‐7382, Santa Cruz Biotechnology), cytochrome c (1:1,000, ADI‐AAM‐175, Enzo Biochem), caspase‐3 (1:1,000, #9662, Cell Signaling Technology), LC3‐I/II (1:500, GTX127375, Gene Tex), p62 (1:500, GTX100685, Gene Tex), Parkin (1:1,000, sc‐137179, Santa Cruz Biotechnology), PINK1 (1:1,000, sc‐32282, Novus Biotechnology), and GAPDH (1:2,000, sc‐137179, Santa Cruz Biotechnology), and then detected using HRP‐conjugated secondary antibodies. The signals were visualized using fluorography with an enhanced detection kit (ECL, GE Healthcare Life Sciences, Buckinghamshire Amersham‐Pharmacia International).

Hsieh C., Li C., Hsu C., Chen H., Chen Y., Liu Y., Liu Y., Kuo H, & Liu P. (2019). Mitochondrial protection by simvastatin against angiotensin II‐mediated heart failure. British Journal of Pharmacology, 176(19), 3791-3804.

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Western Blot Analysis of Autophagy Markers

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Embryos collected at 48 hpf were dechorionated, deyolked, and lysed in radioimmunoprecipitation assay buffer (RIPA buffer) supplemented with 1 mM PMSF, 1 mM sodium fluoride, and 1 mM sodium vanadate (Na₃VO₄). Equal amounts of embryo protein (50 μg) were electrophoresed in 10% SDS-PAGE gel and transferred to nitrocellulose membranes. Western blotting was performed as previously described [80 (link),81 (link)], and the primary antibodies used were: rabbit anti-β-tubulin (#2146, Cell Signaling Technology, Danvers, MA, 1:1000), rabbit anti-LC3 (L7543, Sigma-Aldrich, MI, IT, 1:1000), and rabbit anti-p62 (GTX100685, GeneTex, Irvine, CA, 1:500). The p-value was calculated using GraphPad Prism 6 software (San Diego, CA, USA).

Naef V., Marchese M., Ogi A., Fichi G., Galatolo D., Licitra R., Doccini S., Verri T., Argenton F., Morani F, & Santorelli F.M. (2021). Efficient Neuroprotective Rescue of Sacsin-Related Disease Phenotypes in Zebrafish. International Journal of Molecular Sciences, 22(16), 8401.

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