Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Caco-2 cells labelling BNP with ab92500 at 1/50 dilution (10.8 µg/mL), followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (2 µg/mL) (Green). Confocal image showing cytoplamic staining in Caco-2 cell line. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (2.5 µg/mL) (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (2 µg/mL).

All lanes : Anti-BNP antibody [EPR3735] (ab92500) at 1/2000 dilution

Lane 1 : Human plasma
Lane 2 : Human fetal
heart lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : goat anti-rabbit HRP at 1/2000 dilution

Predicted band size: 15 kDa
Observed band size: 30 kDa why is the actual band size different from the predicted?

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Caco-2 (Human colorectal adenocarcinoma epithelial cell) cells labelling BNP with ab92500 at 1/50 dilution (1µg) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.