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[Santa Cruz] Western (immuno-) Blotting | ||||||||||||||||||
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https://www.scbt.com/resources/protocols/western-immuno-blotting Western (Immuno-) BlottingA. SAMPLE PREPARATIONSample preparation procedures are provided for monolayer cells, suspension cells, and tissue samples. Follow the procedure suited to your needs. Monolayer Cells
Suspension Cells
Tissue Samples
B. ELECTROPHORESIS
C. IMMUNOBLOTTING FOR INDIRECT DETECTION
Indirect Enhanced Chemiluminescence (ECL) Detection
NOTE: If Cruz Marker™ molecular weight standards (sc-2035) are used in the gel, and the membrane has not been incubated with the Cruz Marker™ MW Tag (sc-516729) or Cruz Marker™ MW Tag-HRP (sc-516732), then the Cruz Marker™ compatible mouse IgG binding protein (sc-516102-CM or sc-516132-CM) must be used in order to visualize standards with ECL. NOTE: If Cruz Marker™ molecular weight standards (sc-2035) are used in the gel, and the membrane has been incubated with the Cruz Marker™ MW Tag (sc-516729), then the mouse IgG binding protein m-IgGκ BP-HRP (sc-516102) must be used in order to visualize standards with ECL.
Indirect RGB Fluorescent or Near-Infrared (NIR) DetectionFor RGB Fluorescent or Indirect Near-Infrared (NIR) Western blotting detection, we highly recommend using our kappa or lambda chain mouse IgG binding proteins and a compatible imaging/detection system.
NOTE: For optimal results, use low fluorescence PVDF membranes, such as Immobilon®-FL PVDF: sc-516541, and UltraCruz® Blocking Reagent: sc-516214 for all blocking, primary antibody and mouse IgG binding protein incubations. All RGB Fluorescent or NIR detection steps should be performed in the dark. NOTE: For two-color Western blots using primary antibodies with kappa or lambda light chains, primary antibodies and their corresponding secondary detection reagents may be incubated simultaneously.
NOTE: If Cruz Marker™ molecular weight standards (sc-2035) are used in the gel, and the membrane has been incubated with the Cruz Marker™ MW Tag (sc-516729), then a CFL-conjugated mouse IgGκ binding protein must be used in order to visualize standards.
D. IMMUNOBLOTTING FOR DIRECT DETECTIONDirect Enhanced Chemiluminescence (ECL) DetectionFor direct ECL detection, we recommend the use of SCBT's horseradish peroxidase (HRP)-conjugated primary antibodies.
Direct RGB Fluorescent or Near-Infrared (NIR) DetectionFor Direct RGB Fluorescent or NIR detection, we highly recommend using SCBT's primary antibodies conjugated to Alexa Fluor® dyes, and a compatible imaging/detection system.
NOTE: For optimal results, use low fluorescence PVDF membranes (such as Immobilon®-FL PVDF: sc-516541), and UltraCruz® Blocking Reagent (sc-516214) for both, blocking and dilution of Alexa Fluor® conjugated primary antibodies. All RGB Fluorescent or NIR detection steps should be performed in the dark.
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관리자 | DATE 2022-01-21 15:23:31 | |||||||||||||||||