Nitrocellulose can also be imaged on the Odyssey Imager.Ĭomparison of Millipore Immobilon-FL PVDF and Immobilon-P Western blots on the Odyssey Classic Imaging System. If you normally use PVDF membranes, use the PVDF membrane included in this kit. Ideally, use the IRDye 800CW secondary antibody to detect the low abundant protein and IRDye 680RD to detect the more abundant protein.Ensure primary antibodies are from different species (e.g., rabbit polyclonal and mouse monoclonal).Be careful not to touch the membrane with your hands or gloves.įor one color blots, use IRDye 800CW secondary antibody for detection of the protein. Only handle membranes by the edges with clean forceps. For best results, leave the markers barely visible on the gel, otherwise the marker will be too strong on the Odyssey image.ĭo not write on membranes with regular ink pens or markers, because the ink will fluoresce on Odyssey Imaging Systems. Standard pre-stained blue molecular weight markers can be visualized on Odyssey Imaging Systems.īecause of the sensitivity of Odyssey Imagers, load only 25% of the normal volume. Optional: Prepare a blot using current methods and reagents but detect with secondary antibodies from LI‑COR.One following the LI‑COR protocol for detection with an Odyssey Imaging System.Prepare replicate blots when comparing established methods.Quick Start Guidelines for Western Blot Preparationīlots can be prepared in advance and stored. Immobilon®-FL PVDF Membranes, 4 ( 10 x 10 cm) IRDye 680RD Goat anti-Rabbit Secondary Antibody, 25 µL, 1 mg/mL IRDye® 800CW Goat anti-Mouse Secondary Antibody, 25 µL, 1 mg/mL
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