Ab110727

Manufactured by Abcam

Sourced in United Kingdom

Ab110727 is a primary antibody that recognizes the Glutathione S-Transferase (GST) tag. It can be used for the detection and purification of GST-tagged recombinant proteins.

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3 protocols using ab110727

Protein Expression Analysis in HUVECs

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After extracting the proteins from HUVECs with RIPA (Thermo Fisher Scientific), they were first boiled at 98°C for 5 min to denature. The samples were separated and then transferred to polyvinylidene difluoride (PVDF, Millipore, Bedford, MA, U.S.A.) membranes. The membranes were blocked in 5% non-fat dry milk for 2 h, and incubated with primary antibody of B-cell lymphoma-2 (Bcl-2, 1:1000, ab32124, Abcam, Cambridge, MA, U.S.A.), Bcl-2-associated X (Bax, 1:2000, ab182733, Abcam), cleaved-caspase3 (1:500, ab32042, Abcam), tumor necrosis factor-α (TNFα, 1:1000, ab6671, Abcam), IL-6 (1:1000, 12912S, Cell Signaling Technology, Shanghai, China), IL-8 (1:1000, ab110727, Abcam), IL-1β (1:1000, 12703S, Cell Signaling Technology), TRPM7 (1:1000, ab109438, Abcam) or glyceraldehyde-3-phosphate dehydrogenase (GAPDH, 1:3000, ab8245, Abcam) at 4°C overnight. The membranes were washed and incubated with secondary antibodies for 1 h, and detected by an odyssey infrared imaging system (LI-COR Biosciences, Lincoln, NE, U.S.A.).

Chen X, & Song D. (2020). LPS promotes the progression of sepsis by activation of lncRNA HULC/miR-204-5p/TRPM7 network in HUVECs. Bioscience Reports, 40(6), BSR20200740.

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Immunohistochemical Analysis of Inflammatory Markers

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An IHC analysis was conducted as previously described [10 (link)]. Deparaffinized tissue sections were treated with 3% hydrogen peroxide in methanol for 10 min to remove endogenous peroxidase. Antigen retrieval was carried out with sodium citrate buffer (0.1 M) using the microwave method. The slides were incubated with normal serum to block nonspecific binding and then incubated for 1 h with primary antibodies (diluted 1:100 to 1:200), such as antibodies against IFN-γ (sc-74104; Santa Cruz Biotechnology, Dallas, TX, USA), TNF-α (MBS175453; MyBioSource, San Diego, CA, USA), TGF-β (MBS462142; MyBioSource), IL-8 (ab110727; Abcam, Cambridge, UK), CCL-2 (PAB16617; Abnova, Taipei, Taiwan), CXCL-1 (PAB8798,; Abnova), CXCL-9 (bs-2551R; Bioss, Woburn, MA, USA), CXCL-10 (bs-1502R; Bioss), and CXCL-11 (bs-2552R; Bioss). The slides were incubated for 10 min with biotinylated secondary antibodies (PK-7800; Vector Laboratories, Burlingame, CA, USA) and horseradish peroxidase-conjugated streptavidin. Signals were detected using the 3,3-diaminobenzidine tetrahydrochloride substrate chromogen solution, and the cells were counterstained with Mayer's hematoxylin. In order to evaluate the expression levels, after 5 circles were drawn with equal diameters in separate areas, without overlap, the positive cells were counted from 4 slides per group.

Lee S.Y., Cho J.H., Cho S.S., Bae C.S., Kim G.Y, & Park D.H. (2018). Establishment of a chronic obstructive pulmonary disease mouse model based on the elapsed time after LPS intranasal instillation. Laboratory Animal Research, 34(1), 1-10.

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Intestinal Protein Expression Analysis

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Relative protein levels of CuSOD, MnSOD, GPX1, GPX4, ZO-1, Claudin-1, Occludin, IL-2, IL-8, and IL-10 in the jejunum were determined using western blotting. Colon samples were collected and the protein expression of CuSOD, MnSOD, GPX1, GPX4, IL-2, IL-8 and IL-10 was determined [22 (link)]. The resultant signals were obtained using Quantity One software (Bio-Rad, Hercules, CA, USA). Primary antibodies were used as follows: CuSOD (1:50000; ab51254, Rabbit, Abcam, UK), MnSOD (1:1000; ab68155, Rabbit, Abcam, UK), GPX1 (1:1000; bs-3882R, Rabbit, Bioss, Beijing, China), GPX4 (1:1000; 14432-1-AP, Rabbit, Proteintech, Rosemont, IL, USA), ZO-1 (1:3000; 21773-1-AP, Rabbit, Proteintech, USA), Occludin (1:1000; ab167161, Rabbit, Abcam, UK), Claudin1 (1:500; ab15098, Rabbit, Abcam, UK), IL-2 (1:2000; ab92381, Rabbit, Abcam, UK), IL-8 (1:2000; ab110727, Rabbit, Abcam, UK), IL-10 (1:1000; 20850-1-AP, Rabbit, Proteintech, USA) and Actin (1:5000; 66009-1-Ig, Mouse, Proteintech, USA).

Wang S., Bai M., Xu K., Shao Y., Yang Z., Xiong X., Huang R., Li Y, & Liu H. (2021). Effects of Coated Cysteamine on Oxidative Stress and Inflammation in Weaned Pigs. Animals : an Open Access Journal from MDPI, 11(8), 2217.

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