Immunofluorescence analysis of RAB3A was done on 70% confluent log phase HeLa 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 RAB3A Rabbit Polyclonal Antibody (PA1770) 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 (Product # 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 is a merged image showing Cytoplasmic localization. Panel e is a no primary antibody control. The images were captured at 60X magnification.
|Tested species reactivity||Dog, Hamster, Human, Mouse, Rat|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to residues M(1) A S A T D S R Y G Q K E S S D Q N(18) C of human RAB3A.|
|Storage buffer||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|
|Immunoprecipitation (IP)||Assay dependent|
|Western Blot (WB)||2 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.
PA1-770 detects rab 3A from canine, hamster, human, mouse and rat tissues and cells.
PA1-770 has been successfully used in Western blot and immunoprecipitation procedures. By Western blot, this antibody detects an ~21 kDa protein representing rab 3A from Hepa1 cell lysate.
PA1-770 immunogen is a synthetic peptide corresponding to residues M(1) A S A T D S R Y G Q K E S S D Q N(18) C of human RAB3A. This sequence is completely conserved between human, mouse, and rat rab 3A. PA1-770 immunizing peptide (Cat. # PEP-098) is available for use in neutralization and control experiments.
Recombinant human rab 3A (Cat # RP-770) protein can be purchased for control experiments in Western blot applications.
Rab proteins are low-molecular-weight GTP-binding proteins that form the largest branch of the Ras superfamily of GTPases. Located on the cytoplasmic face of organelles and vesicles, rab proteins are involved in intracellular membrane fusion reactions. Three membrane proteins, synaptosomal associated protein of 25 kDa (SNAP-25), synaptobrevin, and syntaxin, form the core of a ubiquitous membrane fusion machine that interacts with the soluble proteins N-ethylmaleimide-sensitive factor (NSF) and a-SNAP. Rab proteins, in coordination with the core fusion machinery and Munc-18, help to mediate vesicle docking and fusion. There exist over 40 rab proteins that have been described in mammals and the best studied is rab 3A. Rab 3A is found to be abundant in presynaptic nerve terminals and is also found to be crucial in acrosomal exocytosis in human spermatozoa. Abnormal accumulation of rab 3A in the cytoplasm of Purkinje cells has been reported in the prion protein related Creutzfeldt-Jakob disease.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.