Ab28258

Manufactured by Abcam

Ab28258 is a monoclonal antibody targeted towards an undisclosed antigen. It is suitable for use in various immunoassay applications.

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Lab products found in correlation

3 protocols using ab28258

Investigating Inflammatory Signaling Pathways

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IL-1α antibody (SC-7929) was purchased from Santa Cruz Biotechnology. Antibodies directed against phospho-p38 MAPK (4511), p38 MAPK (8690), phospho-JNK (9251), JNK (9252), phospho-ERK (4370), ERK (4695), phosphor-AKT (9611) and AKT (4691) were purchased from Cell Signaling Technology. The antibody for human S100A7 (11141-RP02), soluble recombinant human S100A7 (11141-HNAE), IL17A (12047-HNAE), IL-22 (13059-HNAE), IL-33 (10368-HNAE), IL36γ (10124-HNAE) and TNFα (10602-HNAE) were purchased from Sino Biological Inc. The antibody against β-actin (ab8227), IL-1β (ab2105), RAGE (ab54741) and calpain-1 (ab28258) were purchased from Abcam. Antibody against mS100a7a15 (AP52284) was purchased from Abgent. The inhibitors including SB202190 (S7067), SP600125 (S5567), LY294002 (L9908) and caspase-8 inhibitor (C1230) were purchased from Sigma-Aldrich. PD151746 (S7424) was purchased from Selleck.

Lei H., Li X., Jing B., Xu H, & Wu Y. (2017). Human S100A7 Induces Mature Interleukin1α Expression by RAGE-p38 MAPK-Calpain1 Pathway in Psoriasis. PLoS ONE, 12(1), e0169788.

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Quantification of Neuromuscular Junction Proteins

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Proteins were fractionated by SDS-PAGE (6% or 8% acrylamide; Invitrogen) and transferred to polyvinylidene fluoride membranes. Blots were probed with antibodies to the AChR β-subunit (N8283, RRID:AB_262151; Sigma-Aldrich), Rapsyn (ab156002, RRID:AB_298028; Abcam), MACF1 (gift from R. Liem, Columbia University Medical Center, New York, NY; Chen et al., 2006 (link)), β-Catenin (ab32572, RRID:AB_725966; Abcam), α-Actinin (ab18061, RRID:AB_444218; Abcam), Calpain (ab28258, RRID:AB_725819; Abcam), or MAP1b (ab11266, RRID:AB_297884; Abcam). We used antibodies to the AChR β-subunit to ensure equal loading of proteins in whole-cell lysates and proteins isolated with biotin–α-BGT. We quantified the band intensities with a ChemiDoc imaging system (BioRad), as described previously (Remédio et al., 2016 (link)). The band intensity for each protein was indexed to the band intensity of the AChR β-subunit, and the graphs show the mean values from three separate experiments. The Wilcoxon–Mann–Whitney test was used to determine statistical significance and was conducted using GraphPad Prism 6.0 software.

Oury J., Liu Y., Töpf A., Todorovic S., Hoedt E., Preethish-Kumar V., Neubert T.A., Lin W., Lochmüller H, & Burden S.J. (2019). MACF1 links Rapsyn to microtubule- and actin-binding proteins to maintain neuromuscular synapses. The Journal of Cell Biology, 218(5), 1686-1705.

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Immunoprecipitation of HDAC3, Calpain1, and Calpain2

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Cell lysates were generated by incubating cells with an NP-40 lysis buffer with proteinase inhibitor cocktail on ice and then centrifuged at 10000 g for 10 min. Immunoprecipitation was carried out by incubating the lysate with the corresponding antibodies: anti-HDAC3 (ab47237, Abcam), anti-calpain1 (ab28258, Abcam), anti-calpain2 (ab39165, Abcam) and anti-HA (3724, Cell Signaling Technology) with slow agitation overnight at 4°C. Next, 20 μl PureProteome Protein A/G Mix Magnetic Beads (Merck-Millipore) was added and incubated with slow agitation for 4 h at 4°C. Immune complexes were washed extensively, boiled in SDS-sample buffer and analyzed using SDS-PAGE.

Yang X., Wu Q., Zhang L, & Feng L. (2016). Inhibition of Histone Deacetylase 3 (HDAC3) Mediates Ischemic Preconditioning and Protects Cortical Neurons against Ischemia in Rats. Frontiers in Molecular Neuroscience, 9, 131.

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