Immunofluorescence analysis of GHRL was performed using 70% confluent log phase MKN-45 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with Ghrelin Rabbit Polyclonal Antibody (PA1-1070) at 2ug/ml in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (A27034) a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.
|Tested species reactivity||Human, Mouse, Rat|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to residues F(1) N A P F D V G I K L S G A Q Y Q Q H G R A L(23) of the rat Obestatin gene.|
|Purification||Antigen affinity chromatography|
|Storage buffer||PBS with 1mg/ml BSA|
|Contains||0.05% sodium azide|
|Storage Conditions||-20° C, Avoid Freeze/Thaw Cycles|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||3-5 µg/10^6 cells|
|Immunocytochemistry (ICC)||2 µg/ml|
|Immunofluorescence (IF)||2 µg/ml|
|Western Blot (WB)||1:1000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
PA1-1070 detects Obestatin in mouse and rat samples. This product does not detect endogenous levels of the hormone in rat stomach.
PA1-1070 has been successfully used in Western blot procedures. By Western blot, this antibody detects a ~2.5 kDa protein representing synthetic Obestatin.
The PA1-1070 immunogen is a synthetic peptide corresponding to residues F(1) N A P F D V G I K L S G A Q Y Q Q H G R A L(23) of the rat Obestatin gene. This sequence is conserved in mouse and only one amino acid residue difference in human. This peptide (Cat. # PEP-290) is available for use in neutralization and control experiments.
Obestatin is a peptide hormone. This relatively small protein (~3 kDa) is encoded by the same gene that also encodes ghrelin, also a peptide hormone. The protein produced by that gene breaks into two smaller peptides, ghrelin and obestatin by enzyme cleavage.
Obestatin has been found to suppress appetite whereas ghrelin increases it. Obestatin is produced in the cells that line the stomach and small intestines. Obestatin results in decreased food intake and reduces the gastric emptying activities. It is mainly expressed in the GI tract with higher expressions in the stomach.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.