Worm Lysate from synchronous L3 worm cultures was prepared in Laemmli buffer with 6 M urea with glass beads in a FastPrep machine (MP Biomedicals, Irvine, CA) for 2 × 30 s. Samples were run on a 7.5% SDS-PAGE before transfer onto nitrocellulose membranes (Amersham Protran; 10600003). Membranes were blocked in TBS containing 5% milk for 1 h at RT. First antibody (Monoclonal anti-α-Tubulin clone B-5-1-2, T5168 Sigma-Aldrich 1:20,000 and rabbit polyclonal anti-PTC-3 (Eurogentec, with peptides SASHSSDDESSPAHK and EVRRGPELPKENGLG) 1:500) incubation was done O/N at 4 °C and the secondary HRP-coupled antibodies goat anti-Mouse IgG (H+L) (ThermoFisher scientific; 31430; 1:10,000) or polyclonal HRP-conjugated goat-anti-rabbit IgG (ThermoFisher scientific; 31460; 1:10,000) for 1 h at RT. The blots were developed using WesternBright ECL HRP substrate (K-12045 Advansta) in a Fusion FX7 (Vilber Lourmat) image acquisition system.
Polyclonal hrp conjugated goat anti rabbit igg
Manufactured by Thermo Fisher Scientific
Polyclonal HRP-conjugated goat-anti-rabbit IgG is a secondary antibody conjugated with horseradish peroxidase (HRP). It is designed to detect and bind to rabbit primary antibodies in various immunoassay techniques.
Automatically generated - may contain errors
Lab products found in correlation
Goat anti mouse igg h l, by Thermo Fisher Scientific (1 mentions)
Fastprep, by MP Biomedicals (1 mentions)
Westernbright ecl hrp substrate, by Advansta (1 mentions)
Fusion fx7, by Vilber (1 mentions)
Alexa fluor 488 goat anti rabbit igg h l, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 594 goat anti mouse igg h l cross adsorbed secondary antibodies, by Thermo Fisher Scientific (1 mentions)
Hrp conjugated goat anti mouse igg h l secondary antibody, by Thermo Fisher Scientific (1 mentions)
Ecl prime, by Cytiva (1 mentions)
2 protocols using polyclonal hrp conjugated goat anti rabbit igg
1
Worm Lysate Preparation and Western Blot
2
Antibody Validation for Protein Interactions
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The antibodies used in this study: polyclonal rabbit anti-VIPAS39 (20771-1-AP; proteintech; 1:2000), polyclonal rabbit anti-Rab11-FIP5 (NBP1-81855; Novus Biologicals;1:2,000), polyclonal rabbit anti-EHD1 (NBP2-56035; Novus Biologicals; 1:2,000), polyclonal rabbit anti-rabenosyn-5 (NB300-813; Novus Biologicals; 1:2,000), monoclonal mouse anti-myc (9E10) (1:3,000 for WB and 1:200 for Immunostaining; Sigma-Aldrich; M4439), polyclonal rabbit anti-GFP (TP401; Torrey Pines; 1:3,000 for WB and 1:200 for Immunostaining). For pulldowns Trap beads (Nano bodies) were used. GFP-Trap_A (chromotek, gta-20) for GFP pulldowns and myc-Trap_A (chromotek; yta-20) for myc pulldowns. HRP-conjugated goat anti-Mouse IgG (H+L) Secondary Antibody (Thermo Fisher scientific; 31430; 1:10,000), polyclonal HRP conjugated goat-anti-rabbit IgG (Thermo Fisher scientific; 31460; 1:10,000) were used (incubated for 1 h at RT) to detect bound antibodies with and an ECL system (ECL prime, Amersham, RPN2232). Alexa Fluor 488-goat anti-rabbit IgG (H+L) (Invitrogen; A-11034) and Alexa Fluor 594-goat anti-mouse IgG (H+L) Cross-Adsorbed Secondary Antibodies (Invitrogen; R37121) were used for immunofluorescence.
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