Af6009

Manufactured by Affinity Biosciences

Sourced in United States

The AF6009 is a specialized laboratory equipment used for conducting biochemical and molecular biology experiments. It serves as a reliable tool for various applications, including protein purification, enzyme analysis, and DNA/RNA manipulation. The core function of the AF6009 is to provide a controlled environment for precise experimental procedures, ensuring accurate and reproducible results.

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Lab products found in correlation

3 protocols using af6009

Protein Expression Analysis in Kidney Tissue

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Kidney tissues were homogenized and lysed in RIPA lysis buffer (Beyotime, Shanghai, China). Protein concentrations were determined using a BCA protein assay kit (Thermo Fisher Scientific). Equal amounts of protein (25 μg) were separated using SDS-PAGE and transferred to PVDF membranes (Beyotime). The membranes were blocked with 5% non-fat milk in Tris-buffered saline with Tween 20 (TBST) for 1 h at room temperature and incubated with primary antibodies overnight at 4°C. The primary antibodies used were anti-IRAK2 (1 : 1,000; #DF4782, Affinity, MI, USA), anti-p-NF-κB (1 : 1000; #AF2002, Affinity), anti-NF-κB (1 : 1,000; #AF5006, Affinity), anti-p-IKKβ (1 : 1,000; #AF3010, Affinity), anti-IKKβ (1 : 1,000; #AF6009, Affinity), and anti-GAPDH (1 : 5,000; #AF7021, Affinity). After washing with TBST, membranes were incubated with HRP-conjugated secondary antibodies (1 : 5,000; #S0001, Affinity) for 1 h at room temperature. Protein bands were visualized using an enhanced chemiluminescence kit (Thermo Fisher Scientific) and quantified using the ImageJ software (National Institutes of Health, MD, USA).

Liu J., Xu Y., Cheng S., Wang C, & Zhang Z. (2024). Interleukin-1 receptor-associated kinase 2 promotes inflammatory reactions by activating the nuclear factor kappa-B signaling pathway in diabetic nephropathy. Central-European Journal of Immunology, 48(4), 290-300.

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Comprehensive Protein Extraction and Analysis

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Total protein was extracted from the cortex and neurons by a RIPA lysis buffer containing 1% phenylmethane sulfonyl fluoride and 1% phosphatase inhibitor. The protein content was detected by a BCA Protein Assay Kit (Dingguo Changsheng Biotechnology Co., Ltd., Beijing, China). Western blotting was performed as described previously, and protein bands were developed by an ECL detection system. The following primary antibodies were purchased: anti-CTRP1 (1:1,000, 12209-1-AP; Proteintech, Wuhan, China), anti-β-actin (1:5,000, 20536-1-AP; Proteintech), anti-PERK (1:1,000, AF5304; Affinity), anti-phospho-PERK (Ser555) (1:1,000, AF4499; Affinity), anti-GRP78 (1:1,000, AF53661; Affinity), anti-ATF4 (1:1,000, 10835-1-AP; Proteintech), Bax (1:10,000, 60267-1-AP; Proteintech), Bcl-2 (1:1,000, BF9103; Affinity), eIF2α (1:1,000, AF6087; Affinity), CHOP (1:1,000, AF5280; Affinity), ATF6 (1:1,000, AF6009; Affinity), IRE1α (1:1,000, DF7709; Affinity), phospho-IRE1α (ser724) (1:1,000, AF7150; Affinity), phospho-eIF2α (ser51) (1:1,000, AF3087; Affinity), cleave-caspase 3 (1:1,000, 9661; Cell Signaling Technology, Inc., Danvers, MA, United States), and caspase 12 (1:1,000, 55238-1-AP; Proteintech).

Fei H., Xiang P., Luo W., Tan X., Gu C., Liu M., Chen M., Wang Q, & Yang J. (2021). CTRP1 Attenuates Cerebral Ischemia/Reperfusion Injury via the PERK Signaling Pathway. Frontiers in Cell and Developmental Biology, 9, 700854.

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Investigating Necroptosis Signaling Pathways

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The following commercial antibodies and reagents were used: Necrostatin-1 (S8037, Selleck); Necrostatin 1S (S8641, Selleck); Z-VAD-FMK (S0723, Selleck); TPCA-1(S2824, Selleck); PH-797804 (S2726, Selleck); SP600125 (S1460, Selleck); C-176 (S6575, Selleck); QNZ (S4902, Selleck); RIPK1 (1:1000, 551041, BD Biosciences); RIPK3 (1:1000, NBP1-77299, NOVUS); MLKL (1:2000, AP14272B, Abcepta); pMLKL (1:1000, 37333S, Cell Signaling Technology); Flag (1:1000, 20543-1-AP, Proteintech); caspase3 (1:1000, 19677-1-AP, Proteintech); HMGB1 (1:1000, 10829-1-AP, Proteintech); p65 (1:1000, 66535-1-Ig, Proteintech); GAPDH (1:10,000, 60004-1-Ig, Proteintech); Histone H3 (1:1000, 17168-1-AP, Proteintech);IL6 (1:1000, 66146-1-Ig, Proteintech); CD68 (1:1000, 28058-1-AP, Proteintech); TMEM119 (1:1000, 27585-1-AP, Proteintech); p-p65 (1:200, sc-136548, Santa Cruz); IKK (1:500, AF6009, Affinity); p-IKK (1:500, AF3013, Affinity); IKbα (1:1000, 10268-1-AP, Proteintech); p-IKbα (1:500, AF2002, Affinity); Ccl5 (1:500, AF5151, Affinity); IFNb1 (1:500, DF6471, Affinity); pTau396 (1:1000, 829,001, Biolegend); AT8 (1:1000, MN1020, Thermo); Goat Anti-Mouse-HRP IgG (1:10,000, 115-035-003, Jackson ImmunoResearch); Goat Anti-Rabbit-TRITC IgG (1:200, 111-025-045, Jackson ImmunoResearch); Goat Anti-Rabbit-HRP IgG (1:10,000, 111-005-003, Jackson ImmunoResearch).

Dong Y., Yu H., Li X., Bian K., Zheng Y., Dai M., Feng X., Sun Y., He Y., Yu B., Zhang H., Wu J., Yu X., Wu H, & Kong W. (2022). Hyperphosphorylated tau mediates neuronal death by inducing necroptosis and inflammation in Alzheimer’s disease. Journal of Neuroinflammation, 19, 205.

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