The generation of the anti-CoA antibody (1F10) was described recently [24 (link)]. All common chemicals were obtained from Sigma–Aldrich unless otherwise stated. The following antibodies and dilutions were used: mouse anti-CoA antibody (0.17 μg/ml); rabbit anti-actin antibody (Cell Signaling Technology #4968, 1:2000); rabbit anti-tubulin antibody (Cell Signaling Technology #2148, 1:2000); rabbit anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) antibody (Abcam #ab181602, 1:6000); mouse anti-GSH antibody (Millipore #MAB5310, 1:1000) and rabbit anti-pyruvate dehydrogenase kinase 2 (PDK2) antibody (Abcam #ab68164, 0.5 μg/ml). Primary antibodies were diluted in Odyssey blocking buffer containing 0.01% Tween 20. Secondary antibodies [Alexa Fluor 680 goat anti-mouse IgG H&L (Life Technologies) and IRdye 800 CW goat anti-rabbit IgG H&L (LI-COR Biosciences)] were diluted in Odyssey blocking buffer (1:10 000) containing 0.02% sodium dodecyl sulphate (SDS).
Mab5310
Manufactured by Merck Group
Sourced in United States
MAB5310 is a laboratory equipment product manufactured by Merck Group. It is designed for use in scientific research and analysis. The core function of MAB5310 is to perform specific tasks related to the handling and processing of samples, but a detailed description of its intended use cannot be provided while maintaining an unbiased and factual approach.
Automatically generated - may contain errors
Lab products found in correlation
Rabbit anti actin antibody, by Cell Signaling Technology (2 mentions)
Alexa fluor 680 goat anti mouse igg h l, by Thermo Fisher Scientific (2 mentions)
Irdye 800cw goat anti rabbit igg h l, by LI COR (2 mentions)
Ab92492, by Abcam (1 mentions)
Tris glycine sds sample buffer, by Thermo Fisher Scientific (1 mentions)
4 protocols using mab5310
1
Generation of Anti-CoA Antibody 1F10
2
Detecting Glutathionylation and Na,K-ATPase Isoforms
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Glutathionylation was detected with the anti-glutathione antibody MAB5310 (Millipore). The immunoprecipitation of the glutathionylated proteins was carried out with a monoclonal anti-glutathioine (GSH) antibody (#101-A; Virogen). The Na,K-ATPase α1 isoform was immunoprecipitated with SC-21712 antibody (Santa Cruz Biotech., Dallas, TX) and immunedetected with the sc-28800 antibody (H-300; Santa Cruz Biotech.). All α subunits (α(all)) were immunoprecipitated with sc-28800 and immunodetected with the α5 antibody (Developmental Studies Hybridoma Bank, University of Iowa, Iowa City).
The β1 isoform was immunodetected with a polyclonal antibody generously provided by Dr. P.A Pedersen, University of Copenhagen. The β2 isoform was detected with the polyclonal antibody 06-1711 (Millipore).
The β1 isoform was immunodetected with a polyclonal antibody generously provided by Dr. P.A Pedersen, University of Copenhagen. The β2 isoform was detected with the polyclonal antibody 06-1711 (Millipore).
3
Western Blot Analysis of Hemoglobin S-Glutathionylation
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The effectiveness of hemoglobin S-glutathionylation was assessed using Western blot analysis. After the incubation of hemoglobin with GSSG, a Tris-Glycine SDS sample buffer (“Novex”, Carlsbad, CA, USA) was added without mercaptoethanol. Tris-glycine electrophoresis was performed in 14% of PAAG. Before incubation in a blocking buffer (5% milk), the membranes were fixed with a 5% formalin solution for 40 min to reduce the loss of alpha and beta Hb subunits (12–13 kDa). Primary mouse Anti-Glutathione antibody (“Millipore”, MAB5310 1:1000, Temecula, CA, USA) was used for the detection of S-glutathionylated proteins and rabbit anti-human Recombinant Anti-Hemoglobin subunit beta/ba1 antibody (ab92492, Abcam 1:1000, Germany) for the detection of Hb.
4
Generation and Characterization of Anti-CoA Antibody
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The generation of the anti-CoA antibody (1F10) was described recently [24 (link)]. All common chemicals were obtained from Sigma–Aldrich unless otherwise stated. The following antibodies and dilutions were used: mouse anti-CoA antibody (0.17 µg/ml); rabbit anti-actin antibody (Cell Signaling Technology #4968, 1:2000); rabbit anti-tubulin antibody (Cell Signaling Technology #2148, 1:2000); rabbit anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) antibody (Abcam #ab181602, 1:6000); mouse anti-GSH antibody (Millipore #MAB5310, 1:1000) and rabbit anti-pyruvate dehydrogenase kinase 2 (PDK2) antibody (Abcam #ab68164, 0.5 µg/ml). Primary antibodies were diluted in Odyssey blocking buffer containing 0.01% Tween 20. Secondary antibodies [Alexa Fluor 680 goat anti-mouse IgG H&L (Life Technologies) and IRdye 800 CW goat anti-rabbit IgG H&L (LI-COR Biosciences)] were diluted in Odyssey blocking buffer (1:10 000) containing 0.02% sodium dodecyl sulphate (SDS).
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