Cell Signaling Technology (CST) antibodies are validated utilizing our really useful western blotting protocol, developed and optimized by CST scientists. All knowledge proven on product webpages and datasheets was generated utilizing these protocols. We strongly suggest following our protocols to obtain optimum outcomes with all of our antibodies. Below are just a few examples of the results minor adjustments to the usual protocol could have in your outcomes.
Lysate Preparation
Sonication versus no sonication
Phospho-Histone H3 (Ser10) Antibody #9701
Primary Antibody Dilution Buffer
Milk versus BSA
Phospho-Akt (Ser473) Antibody #9271
Primary Antibody Incubation
Overnight at 4°C versus 2 hours at room temperature
Phospho-Akt (Ser473) Antibody #9271
PKCδ Antibody #2058
Phospho-GSK-3β (Ser9) Antibody #9336
Wash and Dilution Buffer
TBS-T versus PBS-T
Phospho-c-Jun (Ser63) II Antibody #9261
c-Jun (60A8) Rabbit mAb #9165
Ribosomal Protein S3 Antibody #2579
Calnexin (C5C9) Rabbit mAb #2679
Phospho-4E-BP1 (Thr37/46) (236B4) Rabbit mAb #2855
Transfer and Antibody Incubations
Cell Signaling Technology recommends moist switch (as really useful by producer) adopted by one hour blocking and in a single day main antibody incubation at 4˚C.
iBlot® is a dry blotting system that completes switch in 7 minutes. SNAPi.d.™ is a vacuum operated incubation system, which reduces antibody incubation instances to lower than 30 minutes.
Phospho-p38 MAPK (Thr180/Tyr182) (3D7) Rabbit mAb #9215
iBlot® is a trademark of Invitrogen Corporation.
SNAPi.d.™ is a trademark of Millipore Corporation.