2,3,5,4′-Tetrahydroxystilbene glucoside (TSG), purity >98%, was purchased from National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). Sodium azide (NaN3) was obtained from Ameresco (USA). Antibodies against Bcl-2, Bax, β-actin and horseradish peroxidase-conjugated goat anti-rabbit IgG were from Zhongshan Goldenbridge Biotechnology Co., Ltd. (Beijing, China). Annexin V/PI detection apoptotic kit, 2′,7′-dichlorofluorescein diacetate (DCFH-DA) and 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimi- dazolylcarbocyanine iodide (JC-1) were purchased from Beyotime Institute of Biotechnology (Jiangsu, China).
Horseradish peroxidase conjugated goat anti rabbit igg
Manufactured by Zhongshan Golden Bridge Biotechnology
Sourced in China
Horseradish peroxidase-conjugated goat anti-rabbit IgG is a laboratory reagent used in immunoassays. It consists of goat-derived antibodies that bind to rabbit immunoglobulin G (IgG), conjugated to the enzyme horseradish peroxidase. This conjugate can be used to detect and quantify the presence of rabbit IgG in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Pvdf membrane, by Merck Group (2 mentions)
Bca protein assay kit, by Beyotime (2 mentions)
2 7 dichlorofluorescein diacetate dcfh da, by Beyotime (1 mentions)
5 5 6 6 tetrachloro 1 1 3 3 tetraethyl benzimidazolyl carbocyanineiodide jc 1, by Beyotime (1 mentions)
3 3 diaminobenzidine (dab), by Agilent Technologies (1 mentions)
Cellsens dimension, by Olympus (1 mentions)
Anti hsp72 antibody, by Enzo Life Sciences (1 mentions)
Rabbit anti α sma antibody, by Cell Signaling Technology (1 mentions)
Ecl advance western blot detection system, by Bio-Rad (1 mentions)
6 protocols using horseradish peroxidase conjugated goat anti rabbit igg
1
Exploring the Anticancer Potential of TSG
2
Evaluating HBV Infection Models and Therapeutic Targets
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Experiments were performed using the cell models of HepG2.2.15, pHBV1.3-HepG2, and HepG2-N6 cells. Among them, HepG2-N6 cell model was infected with HBV produced in HepG2.2.15 cell culture supernatant and concentrated using PEG8000, and the infection efficiency was verified by western blot. The experiments were divided into five groups: negative control, PSN, PN, SN, and LLN group. Approximately 12 h after the cells were inoculated onto the six-well plates, cells had attained 80% confluency, and 5 nM siRNA-containing nanoparticles or an equivalent amount of nanoparticles were introduced to the cells. Cell supernatants and cells were collected separately after 72 h of incubation, and the expression of HBxAg, pgRNA, HBV DNA and cccDNA were detected using RT-qPCR or qPCR. The primer sequences are shown in Table S1
3
Immunohistochemical Protein Detection Protocol
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IHC was performed as previously described.[15 (link)] Briefly, the 4 μm-thick sections were dewaxed in xylene and gradually rehydrated. Endogenous peroxidases were blocked with 3% hydrogen peroxide. Antigen retrieval was performed by pressure cooking for 2 min in ethylenediaminetetraacetic acid at pH 9.0. After antigen retrieval, the sections were incubated with 10% goat serum for 30 min at room temperature. Subsequently, sections were incubated overnight with rabbit anti-hUTP14a polyclonal antibody (generated in our laboratory) at 4°C and were then incubated with horseradish peroxidase-conjugated goat anti-rabbit IgG (Zhongshan Golden Bridge Biotechnology, Beijing, China) at 37°C for 30 min. Immunocomplexes were visualized using 3,3'-diaminobenzidine (Dako, Carpinteria, CA, USA). Slides were counterstained with light hematoxylin, dehydrated, and cover-slipped.
The histological slides were evaluated by two independent observers blinded to all clinical, pathological, and outcome information, including an experienced pathologist. Any score discrepancy was resolved by discussion to reach a consensus. The specimens were categorized into four groups on the basis of IHC results: negative (−), 0–10% positive cells; faintly positive (+), 10–25% positive cells; moderately positive (++), 25–50% positive cells; and highly positive (+++), and ≥50% positive cells.
The histological slides were evaluated by two independent observers blinded to all clinical, pathological, and outcome information, including an experienced pathologist. Any score discrepancy was resolved by discussion to reach a consensus. The specimens were categorized into four groups on the basis of IHC results: negative (−), 0–10% positive cells; faintly positive (+), 10–25% positive cells; moderately positive (++), 25–50% positive cells; and highly positive (+++), and ≥50% positive cells.
4
Western Blot Analysis of hPER1 Expression
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Cells were lysed in RIPA buffer [50 mmol/L Tris-HCl (pH 7.4), 150 mmol/L NaCl, 0.5% NP-40] for 30 min at 0°C and centrifuged for 15 min (12000 rpm, 4°C). Protein expression was quantified using a BCA Protein Assay Kit according to the manufacturer's instructions (Beyotime, China). The lysates (50 mg protein) were subjected to SDS-PAGE, after which the proteins were transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, USA), which were then blocked with Tris-buffered saline (TBS)-Tween containing 5% non-fat dried milk for 2 h. The membranes were then probed with rabbit polyclonal anti-hPER1 antibody (1:1000, Genetex, USA) and mouse monoclonal anti-hGAPDH antibody (1:3000, Zhongshan Golden-Bridge Biotechnology, China) overnight at 4°C, followed by horseradish peroxidase-conjugated goat anti-rabbit IgG (1:5000, Zhongshan Golden-Bridge Biotechnology, China) at 37°C for 1 h. The precipitated proteins were washed three times in PBS, and an ECL-advance Western Blot Detection System (ChemiDocXRS+, Bio-Rad, USA) was used for detection and photography. The assays were done in triplicate.
5
Immunohistochemical Analysis of Hsp72 and α-SMA
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Immunohistochemistry (IHC) was conducted to assess the expressions of heat shock protein (Hsp) 72 and α-smooth muscle actin (α-SMA) (n = 5 per group). The same sample was cut into 1.5 µm slices. After de-paraffinization, antigen retrieval and endogenous peroxidase blockage. The sections were incubated with rabbit anti-α-SMA antibody (1: 400; Cell Signaling Technology, Cat# 19245S, Beverly, America) and anti-Hsp 72 antibody (1: 100; Enzo Life Sciences, Cat# ADI-SPA-812-F, New York, America) at 4°C overnight, and then incubated with horseradish peroxidase conjugated goat anti-rabbit IgG (Zhongshan Golden Bridge Biotechnology Co. Ltd, Cat# PV-6001, Beijing, China) for 20 min. Staining results were visualized with diaminobenzidine (Zhongshan Golden Bridge Biotechnology Co. Ltd, Cat# ZLI-9079, Beijing, China) and hematoxylin. The images were taken under light microscope (BX51, Olympus, Japan), five high-power fields (400 ×) were chosen and captured (cellSens Dimension, Olympus, Japan). The images were analysis using Image-Pro Plus 6.0 software.
6
Western Blot Analysis of hPER1 Expression
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Exponential phase cells were lysed in RIPA buffer [50 mmol/L Tris- HCl (pH 7.4), 150 mmol/L NaCl, 0.5% NP-40] for 30 min at 0°C. Cell lysates were collected using a cell scraper and centrifuged for 5 min (12,000 rpm, 4°C), and then the supernatants were transferred to new tubes. Protein concentration was quantified using a BCA Protein Assay Kit according to the manufacturer's instructions (Beyotime, Jiangsu, China). The supernatants (50 μg protein) were subjected to SDS-PAGE, after which the proteins were transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, USA), which were then blocked with 5% non-fat dried milk for 1 h. The membranes were then probed with rabbit polyclonal anti-hPER1 antibody (1:1000, Genetex, USA) and mouse monoclonal anti-hGAPDH antibody (1:3000, Zhongshan Golden-Bridge Biotechnology, China) for 2 h at room temperature. The membranes were washed three times in TBS, and then incubated with horseradish peroxidase-conjugated goat anti-rabbit IgG (1:5000, Zhongshan Golden-Bridge Biotechnology, China) at 37°C for 2 h. The precipitated proteins were washed three times in TBS, and an ECL-advance Western Blot Detection System (Bio-Rad, California, USA) was used for detection and photography. The assays were done in triplicate.
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