TrueBlot(R): Anti-IgG HRP

Western Blot of Rabbit TrueBlotR: Anti Rabbit IgG HRP. Lane 1: Rabbit IgG WM reduced (p/n 011 0102) [0.1ug]. Lane 2: Opal Prestained Molecular Weight Marker (p/n MB 210 0500). Lane 3: Rabbit IgG WM non reduced (p/n 011 0102) [0.1ug]. Antibody: Rabbit TrueBlotR: Anti Rabbit IgG HRP at 1.0ug/mL overnight at 4C. Blocking Buffer for Fluorescent Western Blotting (p/n MB 070) for 60mins at RT. Expect: recognizes the Rabbit IgG, only under non reducing condition. Exposure: 0.45sec.

ELISA results of Rabbit TrueBlotR: Anti-Rabbit IgG HRP tested against purified Rabbit IgG protein. Each well was coated in duplicate with 1.0 ug of Rabbit IgG (p/n 011-0102). The starting dilution of antibody was 5μg/ml and the X-axis represents the Log10 of a 3-fold dilution. The titer is 1:450,000. This titration is a 4-parameter curve fit where the IC50 is defined as the titer of the antibody. Assay performed using 3% fish gelatin as blocking buffer and TMB substrate p/n TMBE-1000.

WipB is targeted to lysosomes by its C-terminal domain where it interacts with components of the lysosomal nutrient sensing system (a) HeLa cells expressing GFP-fusion proteins of WipB, WipBD118A, WipB1-364 and WipB365-524. Scale bar, 10？um. (b) HeLa cells expressing GFP-WipB (green) stained using an anti-LAMP-1 antibody (Magenta) after permeabilisation and fixation. Scale bar, 10？um. (c) HeLa cells expressing GFP-WipB (green) were incubated with Lysotracker (red) for 15？min before fixation. Scale bar, 10？um. (d) SDS-PAGE of fractions following co-immunoprecipitation of GFP, GFP-WipB or the indicated GFP-WipB derivatives from HeLa cell lysates using anti-GFP antibody and immunoblotting with anti-v-ATPase A, -v-ATPase B or LAMTOR1 antibodies. A cropped blot is here displayed and the corresponding full-length blot is included in the supplementary information. (e) SDS-PAGE of fractions following co-immunoprecipitation of v-ATPase A, v-ATPase B or LAMTOR1 from lysates of HeLa cells expressing GFP or GFP-WipB and immunoblotting with anti-GFP antibody. A cropped blot is here displayed and the corresponding full-length blot is included in the supplementary information. Figure provided by CiteAb. Source: Sci Rep, PMID: 28842705.

Intersectin-1s (ITSN) interacts via the EH domains with the EHBP1. ECs lysates (250 μg total protein) were subjected to immunoprecipitation with anti-EHBP1 Ab (1 μg), followed by WB with EHBP1 (A) and ITSN1 (B) Abs. EHBP1 Ab brings down the EHBP1 protein as well as ITSN. The 55 kDa immunoreactivity in panel A is cross-reactivity with the IgG heavy chain. The EHBP1 Ab immunoprecipitates the Myc-EHITSN from the stable transfected ECEH-ITSN lysates (B, arrowhead). (C). ECs lysates (250 μg total protein) were subjected to immunoprecipitation with anti-ITSN1 Ab (1 μg), followed by WB with ITSN1 Ab. ITSN1 Ab brings down ITSN protein in both ECEH-ITSN and ECCtrl lysates. The upper ITSN immunoreactivity (190 kDa), belongs to the ITSN-2 long isoform (ITSN-2l). For immunoprecipitation studies (B,C), the rabbit IgG TrueBlot Ab HRP-conjugated which enables detection of immunoblotted target protein bands, without interfering with the immunoprecipitating IgG heavy and light chains has been used. (D) Densitometric analysis of immunoprecipitated ITSN in both ECEH-ITSN and ECCtrl lysates. Data are expressed as ratio of ITSN immunoprecipitated by EHBP1 Ab / ITSN immunoprecipitated by ITSN Ab (D). ？p < 0.05. (E,F). Double anti-ITSN Ab anti-rabbit IgG Alexa Fluor 594-conjugated (E) / anti-EHBP1 Ab ？ anti mouse IgG Alexa Fluor 488-conjugated (F). The merged image reveals significant co-localization ITSN/EHBP1, both in the cytosol and at the plasma membrane (G). (H) The magnification of the boxed area in G, highlights the significant co-localization ITSN/EHBP1 at the plasma membrane level (arrows) and cytosol (arrowheads). Bars: 10 μm (E？G); 5 μm (H); n = 5. Figure provided by CiteAb. Source: Front Physiol, PMID: 30333761.

Intersectin-1s (ITSN) interacts via the EH domains with the EHBP1. ECs lysates (250 μg total protein) were subjected to immunoprecipitation with anti-EHBP1 Ab (1 μg), followed by WB with EHBP1 (A) and ITSN1 (B) Abs. EHBP1 Ab brings down the EHBP1 protein as well as ITSN. The 55 kDa immunoreactivity in panel A is cross-reactivity with the IgG heavy chain. The EHBP1 Ab immunoprecipitates the Myc-EHITSN from the stable transfected ECEH-ITSN lysates (B, arrowhead). (C). ECs lysates (250 μg total protein) were subjected to immunoprecipitation with anti-ITSN1 Ab (1 μg), followed by WB with ITSN1 Ab. ITSN1 Ab brings down ITSN protein in both ECEH-ITSN and ECCtrl lysates. The upper ITSN immunoreactivity (190 kDa), belongs to the ITSN-2 long isoform (ITSN-2l). For immunoprecipitation studies (B,C), the rabbit IgG TrueBlot Ab HRP-conjugated which enables detection of immunoblotted target protein bands, without interfering with the immunoprecipitating IgG heavy and light chains has been used. (D) Densitometric analysis of immunoprecipitated ITSN in both ECEH-ITSN and ECCtrl lysates. Data are expressed as ratio of ITSN immunoprecipitated by EHBP1 Ab / ITSN immunoprecipitated by ITSN Ab (D). *p < 0.05. (E,F). Double anti-ITSN Ab anti-rabbit IgG Alexa Fluor 594-conjugated (E) / anti-EHBP1 Ab ？ anti mouse IgG Alexa Fluor 488-conjugated (F). The merged image reveals significant co-localization ITSN/EHBP1, both in the cytosol and at the plasma membrane (G). (H) The magnification of the boxed area in G, highlights the significant co-localization ITSN/EHBP1 at the plasma membrane level (arrows) and cytosol (arrowheads). Bars: 10 μm (E？G); 5 μm (H); n = 5. Figure provided by CiteAb. Source: Front Physiol, PMID: 30333761.

Rabbit TrueBlotR IP / Western Blot: Jurkat cell lysate (0.5 ml of 1x10e7 cells/ml) was incubated with rabbit anti-human Stat1 and immunoprecipitated using Protein G, Protein A and Anti-Rabbit Ig IP Beads. Precipitate from 5x10e5 cells was subjected to electrophoresis, transferred to a PVDF membrane, and Western blotted with anti-Stat1 using Rabbit TrueBlotR: Anti-Rabbit IgG HRP