Af 410 sp

Manufactured by R&D Systems

The AF-410-SP is a specialized laboratory equipment product manufactured by R&D Systems. It is designed to perform a core function within the research and development environment. The product details and intended applications are presented in a factual and unbiased manner, without any extrapolation or interpretation.

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Lab products found in correlation

P0083, by Beyotime (1 mentions) Interleukin 6 (il 6), by Thermo Fisher Scientific (1 mentions) Tnf α, by Thermo Fisher Scientific (1 mentions) Anti tnf α, by R&D Systems (1 mentions) Mouse tnf alpha elisa kit, by Abcam (1 mentions) Ab 108 c, by R&D Systems (1 mentions) Hepg2 cells, by National Collection of Authenticated Cell Cultures (1 mentions) Hepg2, by R&D Systems (1 mentions) 3t3 l1 preadipocytes, by National Collection of Authenticated Cell Cultures (1 mentions)

6 protocols using af 410 sp

Myocardial Cell Membrane and TNF-α Staining

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The myocardial cell membrane was stained with WGA antibody (ZF0305, Vector Laboratories), diluted with 5% BSA at a ratio of 1:500, incubated for 2 h at room temperature, washed with PBS 1X, and then mounted with DAPI anti-fluorescence quenching mounts, and collected by a microscope.
The paraffin sections of the heart were deparaffinized. According to the instructions of the reagents, used 1% H₂O₂ for 10 min, Used sodium citrate repair solution (P0083, Beyotime) to repair antigen in a microwave oven, and 5% BSA for 30 min. Afterward, incubate with TNF-α primary antibody (AF-410-SP, R&D) at 20 ug/ml at 4°C overnight. After washing with PBS 1X, add anti-goat secondary antibody and incubate for 30 min. SABC reacted for 30 min, washed and stained with DAB staining solution, hematoxylin stained the nucleus, and then mounted the slide and took pictures under a microscope.

Wang Y., Chen T., Yang C., Li Q., Ma M., Xu H., Shi Q., Wang Y., Wang Y, & Liang Q. (2022). Huangqi Guizhi Wuwu Decoction Improves Arthritis and Pathological Damage of Heart and Lung in TNF-Tg Mice. Frontiers in Pharmacology, 13, 871481.

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Cytokine Blocking and Spike-in Assay

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For the cytokine blocking experiment cells were pre-incubated with either phosphate bufferred saline (PBS) or blocking antibodies (anti IL-6, cat. MAB206-SP, R&D 0.15 µg/mL and anti TNF-α, cat. AF-410-SP, R&D 10 µg/mL) for 4 h before the addition of the patients’ serum. Blocking antibodies or PBS were also added at the time of serum addition. Media containing serum and blocking antibodies was refreshed after 48 h and viability and contractility were measured on day 4. For the spike in experiment, IL-6 and TNF- α (Peprotech) were added to media supplemented with serum from control COVID-19 negative patients at the concentration of 100 and 150 pg/mL, respectively.

Colzani M., Bargehr J., Mescia F., Williams E.C., Knight-Schrijver V., Lee J., Summers C., Mohorianu I., Smith K.G., Lyons P.A, & Sinha S. (2023). Proinflammatory cytokines driving cardiotoxicity in COVID-19. Cardiovascular Research, 120(2), 174-187.

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Measuring and Neutralizing TNFα Activity

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Expression of TNFα in conditioned media was assayed using the TNF alpha mouse ELISA kit (Abcam). TNFα activity was neutralized in the myeloid cell-conditioned media using 50 μg/mL mouse TNF-α antibody (R&D Systems; AF-410-SP).

Lim S.Y., Yuzhalin A.E., Gordon-Weeks A.N, & Muschel R.J. (2016). Tumor-infiltrating monocytes/macrophages promote tumor invasion and migration by upregulating S100A8 and S100A9 expression in cancer cells. Oncogene, 35(44), 5735-5745.

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Quantifying TNFα Neutralization in Conditioned Media

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Expression of TNFα in conditioned media was assayed using the TNF alpha mouse enzyme-linked immunosorbent assay kit (Abcam). TNFα activity was neutralized in the myeloid cell-conditioned media using 50 μg/ml mouse TNF-α antibody (R&D Systems; AF-410-SP).

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HDL Cholesterol Efflux in ADAM17-deficient Macrophages

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The cholesterol efflux capacity of HDL (100 μg/ml, 4 hr) was determined in C57BL/6J wild-type and ADAM17-deficient BMDMs radiolabeled with [³H]-cholesterol and incubated with cyclic AMP to induce expression of ABCA1 and ABCG1 and an acyl-coenzyme A:cholesterol acyltransferase inhibitor (2 μg/ml).^{11 (link)} In some experiments, cells were treated with either TNFα (20 ng/ml), a goat anti-mouse blocking TNFα antibody (R&D systems, AF-410-SP) or control IgG (R&D systems, AB-108-C) before measuring cholesterol efflux capacity of HDL. Cholesterol efflux was calculated as the percentage of radiolabeled cholesterol in the medium at the end of the incubation divided by the total radioactivity of the medium and cells.⁴⁴

Kothari V., Tang J., He Y., Kramer F., Kanter J.E, & Bornfeldt K.E. (2021). ADAM17 boosts cholesterol efflux and downstream effects of HDL on inflammatory pathways in macrophages. Arteriosclerosis, thrombosis, and vascular biology, 41(6), 1854-1873.

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Adipocyte-Mediated Regulation of Hepatoma and Melanoma

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H22, B16-F1, human hepatoma HepG2 cells and 3T3-L1 preadipocytes were purchased from China Center for Type Culture Collection (Wuhan, China). The cells were cultured in DMEM media containing penicillin/streptomycin (100 U/100 μg/ml) and 10% fetal bovine serum, and incubated at 37 °C/5% CO₂.
3T3-L1 preadipocytes were differentiated through incubation in complete DMEM containing 10 μg/mL insulin, 0.5 μM dexamethasone, and 0.5 mM 3-isobutyl-1-methylxanthine (IBMX) for 2 days and thereafter in complete DMEM supplemented with 10 μg/mL insulin for next 2 days. Subsequently, cells were maintained in and re-fed every 2 days with culture medium for another 4 days. Differentiation into mature adipocytes was confirmed by Oil Red O staining. Differentiated 3T3-L1 cells were cultured with complete DMEM for 24 h, and supernatants were collected as adipocytes conditional media (adi-CM).
HepG2 or B16-F1 cells were treated with adi-CM for 48 h. In the meantime, anti-TNF-α antibody (R&D system, AF-410-SP) and/or anti-IL-6 antibody (R&D system, MAB406), the inhibitor of STAT3 BP-1-102 (Selleck, S7769) or the inhibitor of NF-κB withaferin A (MCE, HY-N2065) were used at corresponding concentrations.

Li Z., Zhang C., Du J.X., Zhao J., Shi M.T., Jin M.W, & Liu H. (2020). Adipocytes promote tumor progression and induce PD-L1 expression via TNF-α/IL-6 signaling. Cancer Cell International, 20, 179.

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