Cy3 conjugated anti rabbit igg

Anti-ZO-1, anti-occludin, and anti-claudin-3 antibodies were purchased from Invitrogen (Carlsbad, CA). Anti E-Cadherin and anti β-catenin antibodies were purchased from BD Biosciences (Billerica, MA). Horseradish peroxidase (HRP)-conjugated anti-mouse IgG, HRP-conjugated anti-rabbit IgG and anti-β-actin antibodies were obtained from Sigma Aldrich (St. Louis, MO). AlexaFlour-488-conjugated anti-mouse IgG and Cy3-conjugated anti-rabbit IgG were purchased from Molecular Probes (Eugene, OR).

Anti-JNK(pTpY^183/185), anti-c-Src(pY⁴¹⁸), anti-phospho-threonine, anti-ZO-1, anti-occludin, and anti-claudin-4 antibodies were purchased from Invitrogen (Carlsbad, CA). Anti-JNK, anti-Ask1, anti-MKK7 and anti-c-Src antibodies were purchased from Millipore (Billerica, MA). HRP-conjugated anti-mouse IgG, HRP-conjugated anti-rabbit IgG and anti-β-actin antibodies were obtained from Sigma Aldrich. AlexaFlour-488-conjugated anti-mouse IgG and Cy3-conjugated anti-rabbit IgG were purchased from Molecular Probes (Eugene, OR). Anti-E-cadherin, anti-β-catenin, and biotin-conjugated anti-p-Tyr antibodies were purchased from BD Biosciences (Billerica, MA). FITC-conjugated anti-mouse Gr-1 antibodies were from MyBioSource (San Diego, CA).

Cryo-sections of intestine (10 μm thickness) were fixed in acetone methanol mixture (1:1) at −20°C for 2 min and rehydrated in phosphate buffered saline (PBS). Sections were permeabilized with 0.2% Triton X-100 in PBS for 15 min and blocked in 4% non-fat milk in TBST (20mM Tris, pH 7.2 and 150 mM NaCl). It was then incubated for one hour with primary antibodies (mouse monoclonal anti-occludin and rabbit polyclonal anti-ZO-1 antibodies or mouse monoclonal E-cadherin and rabbit polyclonal anti-β-catenin antibodies), followed by incubation for one hour with secondary antibodies (AlexaFluor-488-conjugated anti-mouse IgG and Cy3-conjugated anti-rabbit IgG antibodies from Molecular Probes, Eugene, OR) containing Hoechst 33342. The fluorescence was examined by using a confocal microscope (Zeiss 710) and images from x–y sections (1 μm) were collected using Zen software. Images were stacked using the Image J software (NIH) and processed by Adobe Photoshop (Adobe Systems Inc., San Jose, CA).

Anti-JNK(pTpY^183/185) or pJNK, anti-c-Src(pY⁴¹⁸) or pSrc, anti-ZO-1 and anti-occludin antibodies were purchased from Invitrogen (Carlsbad, CA). Anti-JNK and anti-c-Src antibodies were purchased from Millipore (Billerica, MA). HRP-conjugated anti-mouse IgG, HRP-conjugated anti-rabbit IgG and anti-β-actin antibodies were obtained from Sigma-Aldrich. AlexaFlour-488-conjugated anti-mouse IgG, Cy3-conjugated anti-rabbit IgG, AlexaFluor-488 phalloidin and AlexaFluor-546 phalloidin were purchased from Molecular Probes (Eugene, OR). Anti-E-cadherin and anti-β-catenin antibodies were purchased from BD Biosciences (Billerica, MA).

Hoechst 33342 dye and BODIPY FL-N-(2-aminoethyl) maleimide were purchased from Life Technologies (Grand Island, NY, USA). N-ethylmaleimide and tris (2-carboxyethyl) phosphine were from Sigma-Aldrich (St Louis, MO, USA). All other chemicals were purchased from either Sigma-Aldrich or Thermo Fisher Scientific (Tustin, CA, USA). Anti-ZO-1, anti-occludin, anti-claudin-5 (CLDN-5), and anti-claudin-3 (CLDN-3) antibodies were purchased from Invitrogen (Carlsbad, CA, USA). Anti-E‐Cadherin and anti-β‐catenin antibodies were purchased from BD Biosciences (Billerica, MA, USA). Horseradish peroxidase-conjugated anti-mouse IgG, anti-rabbit IgG, and anti-β-actin antibodies were obtained from Sigma-Aldrich (St Louis, MO, USA). AlexaFlour‐488‐conjugated anti‐mouse IgG and Cy3‐conjugated anti‐rabbit IgG were purchased from Molecular Probes (Eugene, OR, USA). The anti-nrf2 antibody was purchased from Abcam (Cambridge, MA, USA). The anti-cofilinps3 antibody was purchased from Cell Signaling Technology (Danvers, MA, USA).

Anti-ZO-1, anti-occludin, and anti-claudin-3 antibodies were purchased from Invitrogen (Carlsbad, CA). Anti E-Cadherin and anti β-catenin antibodies were purchased from BD Biosciences (Billerica, MA). Horseradish peroxidase (HRP)-conjugated anti-mouse IgG, HRP-conjugated anti-rabbit IgG and anti-β-actin antibodies were obtained from Sigma Aldrich (St. Louis, MO). AlexaFlour-488-conjugated anti-mouse IgG and Cy3-conjugated anti-rabbit IgG were purchased from Molecular Probes (Eugene, OR).

Anti-ZO-1 (rabbit polyclonal), anti-occludin (mouse monoclonal), anti-claudin-2 (mouse monoclonal) and anti-claudin-3 (rabbit polyclonal) antibodies were purchased from Invitrogen (Carlsbad, CA). Rabbit polyclonal anti-toll like receptor-4 (TLR4) antibody was from Santa Cruz Biotechnology, Inc. (Dallas, TX). Anti E-Cadherin (mouse monoclonal) and anti β-catenin (rabbit polyclonal) antibodies were purchased from BD Biosciences (Billerica, MA). Horseradish peroxidase (HRP)-conjugated anti-mouse IgG, HRP-conjugated anti-rabbit IgG and anti-β-actin (mouse monoclonal) antibodies were obtained from Sigma Aldrich. AlexaFlour-488-conjugated anti-mouse IgG and Cy3-conjugated anti-rabbit IgG were purchased from Molecular Probes (Eugene, OR).