Ecl detection system

Manufactured by Vazyme

Sourced in China

The ECL detection system is a laboratory equipment designed for the detection and quantification of chemiluminescent signals. It provides a reliable and sensitive method for analyzing various biological samples, including proteins, nucleic acids, and other biomolecules. The system utilizes the principle of enhanced chemiluminescence (ECL) to generate and capture light signals, enabling accurate and reproducible results.

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Lab products found in correlation

11 protocols using ecl detection system

Silencing Pin1 in Hep3B Cells

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Hep3B cells were seeded in 6‐well plates at 2 × 10⁵ cells/well. DP7‐C and GalNAc‐Pin1 siRNA complex was incubated with the Hep3B cells at doses of 1 μg GalNAc‐Pin1 siRNA. Hep3B cells were harvested 48 hours later for analysis. Total RNA was extracted with a kit (ForeGene). qPCR reactions were performed on a Bio‐Rad CFX96 system.
The western blot results better showed the silencing efficiency of the siRNA. The protein samples were probed with anti‐GAPDH (Cell Signaling Technology, CST), anti‐Pin1 (CST), anti‐Beta Tubulin (CST), anti‐Lamin B (CST), and anti‐Exportin 5 (CST) antibodies. The cells were incubated with primary antibodies at 4°C overnight. Then, the membranes were incubated with secondary antibodies (CST) for 60 minutes and developed using the ECL detection system (Vazyme).

Zhao B., Zhou B., Shi K., Zhang R., Dong C., Xie D., Tang L., Tian Y., Qian Z, & Yang L. (2021). Sustained and targeted delivery of siRNA/DP7‐C nanoparticles from injectable thermosensitive hydrogel for hepatocellular carcinoma therapy. Cancer Science, 112(6), 2481-2492.

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Seasonal Variation of LAMP-1 Expression

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Five testis samples from different turtles in each month (February, May, July, October, and December) in 2014 were homogenized in ice-cold RIPA's buffer (25 mMTris/HCl (pH 7.6), 150 mMNaCl, 1% sodium deoxycholate, 1% Nonidet-P40, 0.1% SDS, 0.05 mM PMSF). The protein concentration was quantified by BCA protein assay (Thermo Fisher Scientific, Rockford, USA). Equal amount of each proteins samples (40 μg/lane) was subjected to electrophoresis on 10% SDS-PAGE and transferred to polyvinylidene di-fluoride (Millipore, Bedford, MA) membranes. After blocking in 5% fat-free dry milk, membranes were incubated with an LAMP-1 antibody (21997-1-AP, Proteintech Group, Chicago, IL) at a dilution of 1:1,000 for 12 h at 4°C. After washing, the membrane was incubated with peroxidase-linked goat anti-rabbit IgG (1:5,000, BS13278, Bioworld Technology Inc., Louis Park, MN) for 2 h. Bound antibodies were detected using the ECL detection system (Vazyme Biotech, China). Immunoreactive bands were quantified with Quantity One software (Bio-Rad Laboratories).

Ahmed N., Yang P., Huang Y., Chen H., Liu T., Wang L., Nabi F., Liu Y, & Chen Q. (2017). Entosis Acts as a Novel Way within Sertoli Cells to Eliminate Spermatozoa in Seminiferous Tubule. Frontiers in Physiology, 8, 361.

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Western Blot Analysis of OVGP Protein

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The samples were homogenized in ice-cold RIPA buffer. The protein concentration was quantified using a BCA protein assay (Thermo Fisher Scientific, Rockford, USA). Equal amounts of protein (30 μg/lane) were subjected to 10% SDS-PAGE and subsequently transferred to polyvinylidene difluoride (PVDF; Millipore, Bedford, MA, USA) membranes. After blocking in 5% fat-free dry milk, the membranes with OVGP and GAPDH (inner reference) were incubated overnight at 4 °C with an anti-OVGP antibody (Santa Cruz Inc., USA) diluted 1:1000 and an anti-GAPDH antibody (Santa Cruz Inc.) separately. After washing, the membrane was incubated with horseradish peroxidase-linked donkey anti-goat IgG (1:3000, Beyotime Biotechnology, CHN) for 2 h. Bound antibodies were detected using an ECL detection system (Vazyme Biotech, China). The immunoreactive bands were quantified using Quantity One software (Bio-Rad Laboratories).

Li Z., Zhang Z., Chen X., Zhou J, & Xiao X.M. (2017). Treatment evaluation of Wharton’s jelly-derived mesenchymal stem cells using a chronic salpingitis model: an animal experiment. Stem Cell Research & Therapy, 8, 232.

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Biliary Exosome Protein Characterization

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Western blot was performed by using equal amounts of protein (10 μg/lane) from the four samples bile, liver, gallbladder, and biliary exosomes. Samples were resuspended in 5 × SDS sample buffer, separated by SDS-PAGE. After the samples were transferred onto nitrocellulose membranes, rabbit anti-CD63 antibody (1:100; Boster Bio-technology, Wuhan, China), rabbit anti-HSP70 antibody (1:00; Huaan Bio-technology, Hangzhou, China), rabbit anti-LAMP-1 antibody (1:100; Huaan Bio-technology, Hangzhou, China), and the β-actin antibody (1:100; Boster Bio-technology, Wuhan, China) were incubated with membranes overnight at 4°C. Finally, the protein band detection was performed using an ECL detection system (Vazyme Bio-technology, Nanjing, China).

Zhu X., Wang S., Tarique I., An T., Yang H., Bai X., Wang X., Chen Q, & Yang P. (2019). Cellular Evidence and Source of Exosomes in the Biliary System of the Chinese Soft-Shelled Turtle, Pelodiscus sinensis. Frontiers in Physiology, 10, 1097.

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Evaluating EMT Markers in GC Tissues

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Proteins were extracted from GC tissues using RIPA lysate (SolarBio Life Science, Beijing, China). Tissue protein lysis products were electrolyzed using 12% SDS/PAGE (Shanghai EpiZyme Biotechnology, Shanghai, China). Thereafter, they were transferred to a polyvinylidene–fluoride membrane (Millipore, Billerica, MA, USA). Immunoblots were visualized by an ECL detection system (Vazyme Biotech Co., Ltd.). Antibodies against GADPH were used as controls. Antibodies against GADPH, N‐cadherin (N‐cad), and vimentin were obtained from Cell Signaling Technology, Danvers, MA, USA. All three antibodies were diluted at 1 : 1000.

Qin X., Chen Y., Ma S., Shen L, & Ju S. (2022). Immune‐related gene TM4SF18 could promote the metastasis of gastric cancer cells and predict the prognosis of gastric cancer patients. Molecular Oncology, 16(22), 4043-4059.

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Protein Expression Analysis of CXCL12

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Fibroblasts were harvested and washed in PBS and then lysed in extraction buffer. Protein contents were determined using a bicin-choninic acid assay kit (Pierce, Rockford, IL, USA). Equal amounts of each protein extract (20 μg per lane) were separated by 10% sodium dodecylsulfate polyacrylamide gel electrophoresis. Following transfer to polyvinylidene fluoride membranes and blocking with 5% nonfat milk diluted in tris buffered saline (TBS), the membranes were incubated with antibodies to CXCL12 (Cat#: AF6612, Beyotime Biotechnology, 1:200) and GAPDH (Cat#: ab37168; Abcam [Cambridge, MA, USA]; 1:2000) overnight at 4°C. GAPDH was used as a loading control. The polyvinylidene fluoride membranes were washed with TBS containing Tween-20 (TBS-T) and then were further incubated with horseradish peroxidase-conjugated secondary antibodies (AS1107; Aspen Biotechnology) at a dilution of 1:10,000 for 1 h at room temperature. Each membrane was then washed again and protein bands were visualized using an enhanced chemiluminescence (ECL) detection system (Vazyme, Nanjing, China). The intensity of each band was quantified using Image-J software (National Institutes of Health, Bethesda, MD, USA) and was normalized against GAPDH.

Liao Z.K., Hu S.H., Han B.Y., Qiu X., Jiang S, & Lei T.C. (2021). Pro-pigmentary action of 5-fluorouracil through the stimulated secretion of CXCL12 by dermal fibroblasts. Chinese Medical Journal, 134(20), 2475-2482.

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Quantifying Androgen Receptor Expression in Pelodiscus sinensis

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The samples were homogenized in ice-cold RIPA buffer (25 mM Tris/HCl (pH 7.6), 150 mM NaCl, 1% sodium deoxycholate, 1% Nonidet-P40, 0.1% SDS, and 0.05 mM PMSF). The protein concentration was quantified using a BCA protein assay (Thermo Fisher Scientific, Rockford, USA). Equal amounts of protein (40 μg/lane) were subjected to 8% SDS-PAGE and subsequently transferred to polyvinylidene di-fluoride (PVDF) (Millipore, Bedford, MA) membranes. After blocking in 5% fat-free dry milk, the membranes were incubated overnight at 4 °C with an anti-AR antibody (ab198394, Abcam Inc., Cambridge, MA, USA) diluted 1:1000. After washing, the membrane was incubated with peroxidase-linked goat anti-rabbit IgG (1:5000, BS13278, Bioworld Technology Inc., Louis Park, MN) for 2 h. Bound antibodies were detected using an ECL detection system (Vazyme Biotech, China). The immunoreactive bands were quantified using Quantity One software (Bio-Rad Laboratories). The validity of AR antibody in P. sinensis has been detected by negative and positive control analysis.

Liu T., Chu X., Huang Y., Yang P., Li Q., Hu L., Chen H, & Chen Q. (2016). Androgen-related sperm storage in oviduct of Chinese Soft-Shelled Turtle in vivo during annual cycle. Scientific Reports, 6, 20456.

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Testis Protein Extraction and Western Blot

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Samples of the testis in each group were homogenized in ice-cold RIPA buffer (25 mM Tris/HCl (pH 7.6), 150 mM NaCl, 1% sodium deoxycholate, 1% Nonidet-P40, 0.1% SDS, 0.05 mM PMSF), and centrifuged at 15,000 g for 10 min at 4°C. Then, the total protein concentration was determined with a BCA protein assay (Santa Cruz, sc-202389). Samples (40 μg protein per lane) were subjected to electrophoresis on a 10% SDS-PAGE gel and then transferred onto PVDF membranes (Millipore, ISEQ00010). After nonspecific blocking in 5% nonfat milk, the membranes were incubated with an anti-LC3B (1:1000 dilution) antibody (Abcam, ab48394) overnight at 4°C. After washing with TBST, the membranes were incubated with peroxidase-linked goat anti-rabbit IgG (1:5000, Bioworld Technology Inc., BS13278) for 2 h. Following incubation, the bound antibodies were visualized by using the ECL detection system (Vazyme Biotech, E411-04). Immunoreactive bands were quantified with Quantity One software (Bio-Rad Laboratories).

Huang Y., Yang P., Liu T., Chen H., Chu X., Ahmad N., Zhang Q., Li Q., Hu L., Liu Y, & Chen Q. (2016). Subcellular Evidence for Biogenesis of Autophagosomal Membrane during Spermiogenesis In vivo. Frontiers in Physiology, 7, 470.

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Western Blot Analysis of LC3B in Oviduct

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Western blot was performed as described previously20 (link) using equal amounts of protein (40 μg/lane) from the oviduct. Anti-LC3B antibody (ab48394, Abcam Inc., Cambridge, MA, USA) was used at a concentration of 2 μg/ml, and protein band detection was performed using an ECL detection system (Vazyme Biotech, China). The validity of the anti-LC3B antibody in P. sinensis was determined by negative and positive control analysis (Supplementary Fig. S1).

Liu T., Yang P., Chen H., Huang Y., Liu Y., Waqas Y., Ahmed N., Chu X, & Chen Q. (2016). Global analysis of differential gene expression related to long-term sperm storage in oviduct of Chinese Soft-Shelled Turtle Pelodiscus sinensis. Scientific Reports, 6, 33296.

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Western Blot Analysis of Protein Lysates

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Cells were washed in ice-cold PBS, then lysed and harvested in Radio-Immunoprecipitation Assay buffer (RIPA buffer) in the presence of protease inhibitor cocktail. Protein concentrations were measured using a BCA Kit (Beyotime, Shanghai, China), and separated by SDS-PAGE on 12.5% gels (Beyotime, Shanghai, China), then electroblotted onto polyvinylidene fluoride (PVDF) membrane. The membranes were incubated with the primary antibody overnight, and the blots were incubated with horseradish peroxidase (HRP)-conjugated secondary antibody for 45 min at room temperature. Finally, the blots were exposed using an ECL detection system (Vazyme, Nanjing, China), and Western blotting bands were quantified according to intensity using ImageJ software.

Zhang X.Z., Tian W.J., Wang J., You J.L, & Wang X.J. (2022). Death Receptor DR5 as a Proviral Factor for Viral Entry and Replication of Coronavirus PEDV. Viruses, 14(12), 2724.

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