![]() ![]() Incubate with ECL reagent for 1 min, cover with Saran wrap (to remove excess solution from the surface), and expose to X-ray film in a dark room. Wash three times with TBS-T (1 for 15 min, 2 for 5 min) and then once with TBS (5 min). For optimal antibody dilution, follow the manufacturer's recommendations. Incubate for 30 min at room temperature with the secondary antibody conjugated to HRP. Incubate for 30 min at room temperature with primary antibody (purified antibody 0.1-10 µg/mL, antiserum 1:1,000-100,000, hybridoma supernatant 1:100-10,000) in BSA/TBS-T. Apply slowly, minimizing the area of solution penetration (usually 2-4 mm in diameter).īlock non-specific sites in 10 cm Petri dishes with 5% BSA in TBS-T by immersion (30 min to 1 h at room temperature). Use a narrow-mouthed pipette tip and spot 2 µL of sample onto the nitrocellulose membrane at the center of the grid. Draw a grid with a pencil to indicate the area where you want to make the blot. TBS-T is a TBS solution containing 0.05% Tween 20.īSA/TBS-T is a TBS-T solution containing 0.1% BSA. TBS is made of 20 mm Tris-HCl and 150 mm NaCl with a pH of 7.5. Creative Biolabs can develop specific antibodies for you to use to perform dot blotting assays. If you have a purified protein and a specific antibody, you can use the dot blotting method to make semi-quantitative estimates of protein concentrations in crude preparations such as culture supernatants. Please keep in mind that Creative Biolabs cannot guarantee specific results for dot blotting assays.ĭot blotting is a technique used to detect, analyze, and identify proteins, which is similar to western blotting, but differs in that protein samples are not separated by electrophoresis, but are spotted directly onto a membrane or paper substrate by means of a circular template. This procedure should be used as a guideline only. With strict regulation and effective execution, we are dedicated to providing the most valuable solutions to complete your projects. Cell-based Membrane Protein Expression and PurificationĬreative Biolabs is offering the most comprehensive services for antibody development projects.Cell-free Membrane Protein Expression and Purification.Membrane Protein Solubilization and Stabilization.Manufacturability Assessment and Optimization.SIAT ® Immunogenicity Assessment Services.CreMap™ Epitope Mapping & Discovery Services.Dual-Polarization Interferometry (DPI) Services.Differential Scanning Calorimetry (DSC) Services.Isothermal Titration Calorimetry (ITC) Services.De Novo Human/Humanized Antibody Sequencing Service.Magic™ Human Antibody Production Using Transgenic Mice.Bovine Ultralong CDR3 Antibody Humanization.Antibody Off-target Identification and Specificity Profiling Service.Biomolecular Interaction Analysis Services.Epitope Prediction for the Identification of Antibody Epitopes.Epitope Mapping by X-ray Crystallography.Phage Display & Antibody Library Services. ![]()
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