Vacuum Transport
SNAP i.d.® 2.0 System: Decrease immunodetection time by 80%
compared to traditional Western blotting
This novel method allows you to optimize your Western blotting conditions in record time for maximum results. The SNAP i.d.® 2.0 system is compatible with fluorescent, chemiluminescent or chromogenic detection. Moreover, the sequence of steps required to process a Western blot with the SNAP i.d.® 2.0 system is identical to that used in traditional immunodetection. Because it uses a vacuum to actively drive reagents through the membrane, blocking and washing steps are faster and more thorough.
Advantages of the SNAP i.d.® 2.0 System’s Vacuum Transport Feature
Traditional immunodetection relies on the slow diffusion of reagents into and out of the blot, leading to long incubation times and possible high background. The SNAP i.d.® 2.0 system actively pulls the antibodies through the membrane for maximum interaction with the antigens without a residual high background. The system increases local antibody concentrations at binding sites by using vacuum filtration as well as decreased antibody volumes, driving the antibody-antigen binding reaction forward and shortening incubation times. Vacuum pulls any residual, unbound antibody out of the membrane, lowering background signal.
- Draws reagents through blotting membrane
- Minimizes over-blocking
- Membranes are thoroughly flushed instead of just rinsed
- Reduced incubation times
Low concentrations of blocking reagent with SNAP i.d. improve quality.
Non-Fat Dry Milk (NFDM) is a blocking solution commonly used in Western blotting; however, its high blocking capacity may compromise the protein signal. To demonstrate this, a two-fold dilution series of rat liver lysate (12 µg in lane 1 to 0.09 µg in lane 8) was resolved with SDS-PAGE prior to blotting and immunodetection (the primary antibody was mouse anti-GAPDH; the secondary antibody was HRP conjugated goat anti-mouse). Blot A used a traditional immunodetection protocol (block for 1 hour in 5% NFDM, incubate in primary or secondary antibody at 1:50,000 for 1 hour, wash three times following incubations). Blots B and C were assembled in SNAP i.d blot holders and blocked for 20 s with either 0.5 or 0.05% NFDM, respectively. The blots were incubated for 10 m with anti-GAPDH (1:13,000), washed immediately and incubated for 10 m with HRP goat anti-mouse (1:10,000). Results show an increase in sensitivity with a decrease in milk concentration.
Improve your signal!
Use low concentrations of blocking reagents with the SNAP i.d. system to improve quality.
Non-Fat Dry Milk (NFDM) is an efficient blocking solution commonly
used in western blotting; however, its high blocking capacity may
compromise the protein signal. To demonstrate this, a two-fold
dilution series of rat liver lysate (12 μg in lane 1 to 0.09 μg in lane 8)
was resolved with SDS-PAGE prior to blotting and immunodetection.
(The primary antibody was mouse anti-Glyceraldehyde-3-Phosphate
Dehydrogenase (GAPDH); the secondary antibody was HRP-conjugated
goat anti-mouse). Blot A used a standard immunodetection protocol
(block for 1 hour in 5% NFDM, incubate in primary (1:40,000) or
secondary antibody (1:50,000) for one hour, wash three times
following incubations).
Blot B, C and D were assembled in SNAP i.d blot holders and blocked
for 20 seconds with either 0.5, 0.1 or 0.05% NFDM respectively.
The blots were incubated for 10 minutes with anti-GAPDH (1:13,000),
washed immediately and incubated for 10 minutes with HRP goat
anti-mouse (1:10,000). Results show an increase in sensitivity with a
decrease in milk concentration.
