b'Restore Western Blot Stripping BufferThermo Scientific Restore Western Blot Stripping Buffer safely and effectively removes primary and secondary antibodies from nitrocellulose and PVDF membranes to allow chemiluminescent western blots to be reprobed. By stripping and reprobing, there is no need to waste rare or costly samples by running multiple gels in order to probe for different targets (Figures 4850). The procedure with Restore stripping buffer only takes 15 to 30 minutes, depending on the affinity of the primary antibody.A PDI C Actin E Hsp90, PDI, -tubulinFeatures: 10070 Saves timeno need to rerun gels and blots 5540 Saves costly samplereprobe the membrane A using theC Actin E Hsp90, PDI, -tubulin HspS9 t r0Ai,pc P ttiDensI,t2-tubulin Hsp90, PDI, -tubulinPDI A PDI CBA EDC E100 AcSt ti nrPipD tIest 1same target sample 70 100 10055 70 7040 55 55 Effectiveformulation is more efficient at stripping40 40B A DC E Hsp90, PDI, -tubulinantibodies than homemade buffers St r iPpD tIest 1 BSt r iApc tteinst 2 D B D Strip test 2100 Strip test 1 Stri p S tterispt t2est 17055 Gentledoes not damage the target protein during40stripping, allowing efficient reprobingB Strip test 1 D Strip test 2Figure 48. Stripping and reprobing blots for similar molecular weightOdor-freeno mercaptans means no acrid odors targets with Restore Western Blot Stripping Buffer. A431 cell lysate was diluted to 125 g/mL in electrophoresis reducing sample buffer, and 2-fold serial dilutions were made. 10 L of each dilution (1,250 ng toEconomicalless expensive than other commercial39 ng of total protein) was separated by SDS-PAGE and transferred to a stripping buffers 0.45 m nitrocellulose membrane. The membrane was blocked with 5% nonfat dry milk in 1X Thermo Scientific Pierce PBS Tween 20 Buffer and analyzed by western blot using SuperSignal West Dura Extended Duration Substrate and imaged. (A) The first target was detected by probing with Invitrogen PDI Monoclonal Antibody at 0.33 g/mL, followed by Invitrogen Goat AntiMouse IgG HRP Conjugate at 6.7 ng/mL and imaged. (B) Next, the blot was stripped in Restore Western Blot Stripping Buffer for 15 minutes at 37C, washed in 1X Pierce PBS Tween 20 Buffer, View how-to video: incubated with substrate, and imaged to check for stripping efficiency. (C) The second target was detected by reblocking the membrane and probing with Invitrogen Actin Monoclonal Antibody at 0.5 g/mL, followed by the antimouse IgG HRP conjugate at 6.7 ng/mL and imaged. (D) The blot was stripped again and then (E) probed for multiple targets (-tubulin at 0.2 g/mL, PDI at 0.33 g/mL, and hsp90 at 0.14 g/mL), and imaged as described above.71'