We also found that RS enhances A�� generation by increasing selleck chem the protein level of APP. To further prove this we compared the effect of HLJDT with and without RS (modified HLJDT, or HLJDT-M) on APP processing and A�� load in N2a-SwedAPP cells. We meticulously evaluated the effect of each individual herb of HJLDT on the levels of APP, sAPP�� and sAPP�� in order to determine which herb is responsible for increasing APP and A��. Materials and Methods Chemicals and Reagents Dulbecco’s modified Eagle’s medium (DMEM), fetal bovine serum (FBS), penicillin, streptomycin and G418 for cell culture were bought from Invitrogen (Carlsbad, CA, USA). Polyvinylidene Fluoride (PVDF) membrane was obtained from Hybond-P, GE Healthcare BioSciences (Piscataway, NJ, USA).

Enhanced chemiluminescence (ECL) reagent was purchased from Thermo Scientific (Rockford, IL, USA). Tetramethylbenzidine (TMB) was purchased from BD Biosciences (Sparks, MD, USA), while analytical grade reagents (including ethanol and methanol) were from Sigma�CAldrich (St. Louis, MO, USA) unless otherwise indicated. Berberine, baicalin and baicalein were purchased from Sigma-Aldrich. Palmatine and geniposide were purchased from Aktin Chemicals (Chengdu, China). Monoclonal ��-actin antibody was purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Rabbit polyclonal CT15 antibody against the C-terminus of APP was a gift from Prof. Edward Koo (University of California, San Diego, La Jolla, CA, USA). Amino-terminus anti-A��1�C17 monoclonal antibody (6E10) and a biotinylated A��17�C24 (4G8) monoclonal antibody wereordered from Covance (Princeton, NJ, USA).

Human anti-sAPP�� Swedish (sAPP��-sw) monoclonal antibody (clone 6A1) was provided by IBL (Japan). Anti-pAPPThr668 polyclonal antibody was from Cell Signaling (Danvers, MA, USA). Carboxy-terminus biotinylated anti-amyloid bG4(1�C40)-5C3 (specific to a peptide corresponding to A��40) and bA4(1�C42)-8G7 (specific to a peptide corresponding to A��42) antibodies were purchased from Nanotools (Teningen, Germany). The streptavidin-conjugated horseradish peroxidase (HRP) was purchased from DAKO (Carpinteria, CA, USA). A��40 and A��42 peptides were provided by Invitrogen (Carlsbad, CA, USA) and California Peptide (Napa, CA, USA), respectively. Plant Extraction RC, RS, CP and FG were purchased from the Hong Kong Baptist University Mr. & Mrs. Chan Hon Yin Chinese Medicine Specialty Clinic and Good Clinical Practice GSK-3 Centre and from a local Chinese medicine pharmacy. All herbs were identified and authenticated by Prof. Zhong-Zhen Zhao from the School of Chinese Medicine, Hong Kong Baptist University, Hong Kong. Dry materials of the plants were ground into powder.