WB Quickly 1 min blocking buffer
No: F1BLOK500P
Size: 500ml
Store at 2~8 ℃
Before use this WB 1min blocking buffer: Avoid touching the membrane with your fingers; use tweezers instead. Oils and proteins on the fingers will block efficient transfer and create dirty blots. After sandwiching the gel and membrane between paper, air bubbles between the gel and membrane can be removed by rolling them out with a pipet or 15 ml tube, or by assembling the sandwich in a dish of transfer buffer to prevent formation of bubbles in the first place. Wear gloves! Make sure the paper and membrane are cut to the same size as the gel. Large overhangs may prevent a current from passing through the membrane in semi-dry transfers. Chicken antibodies tend to bind to PVDF and other nylon-based membranes, leading to high background. Switching to a nitrocellulose membrane should help reduce background staining 1. Immerse the PVDF or NC paper with WB 1min blocking buffer (Please note that incubation with membrane with WB 1min blocking buffer for 1-3 mins is suitable, however, the prolonged incubation will result in the high backgroud). Blocking the membrane: Blocking the membrane prevents non-specific background binding of the primary and/or secondary antibodies to the membrane (which has a high capacity for binding proteins and therefore antibodies). 2. Incubation with primary antibody: Dilute the antibody in WB 1min blocking buffer (provided) at the suggested dilution. Incubate your membrane for a few hours to overnight (depend on the binding ability of your antibody for the protein). 3. Wash the membrane several times in WB 1min blocking buffer while agitating, 5 minutes or more per wash, to remove residual primary antibody. 4. Incubation with secondary antibody: Dilute the antibody in WB 1min blocking buffer (provided) at the suggested dilution. Incubate your membrane for 1-2 hours (for the secondary antibody, 2 hours incubation is the longest time!!). 5. Wash the membrane several times in WB 1min blocking buffer while agitating, 5 minutes or more per wash, to remove residual secondary antibody. 6. Add colormetric/ Chemiluminescent substrates to perform color development methods. Recommended storage temperature: 2 ~ 8℃ |