Anti-BiP | Lumenal-binding protein (chicken antibody)

Application example

5 µg of total protein from  A.thaliana (1), H. vulgare (2),  Z.mays (3),  S. oleracea (4), extracted with Agrisera PEB extraction buffer (AS08 300) were separated on  4-12% SDS-PAGE and blotted 1h to PVDF. Blots were blocked immediately following transfer in  for 1h at room temperature with agitation. Blots were incubated in the primary antibody at a dilution of 1: 10 000 for 1h at room temperature with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, then washed once for 15 min and 3 times for 5 min in TBS-T at room temperature with agitation. Blots were incubated in secondary antibody (anti-hen IgY horse radish peroxidase conjugated, from  Agrisera AS09 603) diluted to 1:50 000  for 1h at room temperature with agitation. The blots were washed as above and developed for 5 min with ECL  detection reagent according to the manufacturers instructions.  Exposure time was 5 seconds.

immunofluorescence

BiP localization in 5 days old Arabidopsis thaliana roots. BiP signal shown in green, DAPI in blue. The material has been fixed in para-formaldehyde for 30 minutes. Tissue cleaning has been performed before immunolocalization. Chicken anti-BiP primary antibody was diluted in 1: 1000 and DyLight®488 conjugated goat anti-chicken secondary antibody AS09 622 (green color) was diluted in 1: 1000. Co-staining with DAPI visualized nucleus (blue color).  Scale bar – 10 µm.

BiP2 (Binding immunoglobulin protein) is localized in endoplasmic reticulum lumen (ER) and plays a role in protein assembly inside ER.  BiP protein is abundant under all growth conditions but its synthesis can increase under conditions that lead to the accumulation of unfolded polypeptides in endoplasmic reticulum (ER). Alternative name: AtBP2