Immunofluorescence analysis of Rabphilin 3A was performed using 70% confluent log phase RSC96 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 RPH3A Rabbit Polyclonal Antibody (PA1774) at 2µg/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) at 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 cytosolic localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.
|Tested species reactivity||Rat|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to residues M(1) T D T V V N R W M Y P G D G P L Q(18) C of rat Rabphilin 3A.|
|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 *|
|Immunocytochemistry (ICC)||2 µg/ml|
|Immunofluorescence (IF)||2 µg/ml|
|Western Blot (WB)||2-4 µg/ml|
* 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.
PA1-774 detects rabphilin 3A from rat tissues.
PA1-774 has been successfully used in Western blot procedures. By Western blot, this antibody detects an ~75 kDa protein representing rabphilin 3A from rat brain whole protein extract.
PA1-774 immunogen is a synthetic peptide corresponding to residues M(1) T D T V V N R W M Y P G D G P L Q(18) C of rat Rabphilin 3A. This sequence is completely conserved between rat and mouse rabphilin 3A. PA1-774 immunizing peptide (Cat. # PEP-102) is available for use in neutralization and control experiments.
Rabphilin 3A, a target protein for the small GTPase binding protein rab 3A, plays a role in regulated secretion, particularly in neurotransmitter release. Double labeling of rab 3A and rabphilin 3A shows the two proteins to be co-localized in nerve terminals of the spinal cord and motor endplates. Rabphilin binds selectively to the GTP bound form of rab 3A as well as of rab 3c. Studies suggest that rabphilin 3A phosphorylation by CaM kinase II modulates neurotransmitter release.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.