Goat anti mouse igg

Blood donations were from healthy volunteers who provided written informed consent. The pMH3 vector and CHO-S cells were purchased from AmProtein (Hangzhou, China). Restriction enzymes, T4 DNA ligase, and Taq DNA polymerase were obtained from New England BioLabs (Beverly, MA, USA). Rabbit anti-human VWF polyclonal antibody and rabbit anti-human VWF-HRP antibody were purchased from Dako Cytomation (Glostrup, Denmark). Ristocetin, tetramethylbenizidine (TMB), and collagen type III from human placenta were purchased from Sigma (St. Louis, MO, USA). Goat anti-mouse IgG and Goat anti-mouse IgG labeled with horseradish peroxidase (HRP) were purchased from Beckman-Coulter (Brea, CA, USA). SZ-123, a murine anti-human VWF A3 domain mAb (IgG1) was produced by standard hybridoma technology in our laboratory as described previously [22 (link)]. SZ-34, a murine mAb that binds to the VWF A2 domain but does not inhibit its function, was produced by standard hybridoma technology in our laboratory [25 (link)].

In electrophoresis for protein separation, each well contained a respective sample with 50 µg of proteins. Proteins were transferred onto PVDF (polyvinylidene fluoride) membrane through the wet-transfer method. A mouse anti-HSP70 (clone N27F3-4) monoclonal antibody was purchased from Enzo Life Sciences (New York, USA). A mouse anti-GAPDH (clone 1D4) monoclonal antibody was purchased from Novus Biologicals (Denver, USA). Horseradish peroxidase conjugated secondary antibodies; goat anti-mouse IgG (Beckman Coulter, Brea, CA, USA) was used for to identify the bands reactive to the primary antibodies through an enhanced chemiluminescence reagent (Pierce Biotechnology Inc., Rockford, IL, USA). Primary and secondary antibodies were incubated with membranes at 1:1000 and 1:5000 dilation, respectively. Signaling was quantified by the luminescence image analyzer ImageQuant LAS 4000 (GE Healthcare Life Sciences).