Affinipure goat anti mouse igg h l antibody

The pancreatic isolated islets were fixed and paraffin embedded. Tissue sections
were incubated with primary insulin antibody (Abcam, Cambridge, MA, USA), along
with secondary AffiniPure Goat Anti-Mouse IgG H&L antibody (Jackson
Immunoresearch Laboratories and Molecular Probes, West Grove, PA, USA).
Counterstaining was performed with DAPI (Vector, Burlingame, CA, USA).
Pannoramic MIDI and Pannoramic Viewer (3DHistech, Budapest, Hungary) were used
to scan stained slides and capture images. All the pictures were quantified and
analyzed using Image-Pro Plus software.

Immunofluorescence was used to detect cell-surface expression of vWF, which is an endothelial cell-specific gene. Cells (1×10³ cells/well) were seeded on coverslips for 3 days, and then fixed in 1% paraformaldehyde at 4°C for at least 10 min. After being washed twice (2 min each time) with PBS, cells were permeabilized with 0.1% Triton X-100 (Sigma-Aldrich; Merck KGaA) four times, 5 min each time. Membranes were blocked with goat serum (cat no. SL038; dilution 1:50; Beijing Solarbio Science & Technology Co., Ltd., Beijing, China) at 37°C for 10 min. Subsequently, cells were incubated with 100 µl mouse anti-human vWF antibody (cat no. sc-516102; dilution 1:50; Santa Cruz Biotechnology, Inc., Dallas, TX, USA) for 1 h at 37°C. After washing three times (2 min each time) with PBS, the cells were incubated with the secondary antibody, FITC-conjugated AffiniPure goat anti-mouse IgG (H+L) antibody (cat no. 127-095-099; dilution 1:50; Jackson Laboratory, Ben Harbor, ME, USA), at 37°C for 1 h. Nuclear counterstaining was performed with 1.5 µg/ml DAPI (Biotium, Inc., Hayward, CA, USA) for 30 min at 4°C. Immunofluorescence was observed under a fluorescence microscope. Four replicates were performed.