Immunofluorescent analysis of beta-2 Adaptin was performed using 70% confluent log phase SH-SY5Y 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 beta-2 Adaptin Rabbit Polyclonal Antibody (PA1-1066) 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 punctate cytoplasmic localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.
|Tested species reactivity||Bovine, Human, Mouse, Rat|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to residues I(588) H R K H L P I H H G S T D A G D S P(606) of human beta Adaptin 2.|
|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)||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.
PA1-1066 detects beta Adaptin 2 from human, mouse, rat, and bovine samples.
PA1-1066 has been successfully used in Western blot procedures.
The PA1-1066 immunogen is a synthetic peptide corresponding to residues I(588) H R K H L P I H H G S T D A G D S P(606) of human beta Adaptin 2.
Clathrin-mediated endocytosis is the pathway by which many receptors for nutrients and hormones are internalized to be recycled or down-regulated. During formation of clathrin coated membranes, clathrin co-assembles with heterotetrameric molecules known as assembly polypeptides (APs) or adaptors which form a layer of protein coat between the clathrin lattice and the membrane.
There are two characterized adaptors AP1 and AP2. AP1 is associated with clathrin coated vesicles at the trans-Golgi network and AP2 is associated with the endocytic clathrin coated vesicles at the plasma membrane and has been shown to specifically interact with Shc and EGF receptor. AP2 is composed of four subunits, two separate ~100 kDa gene products with similar domain structures (alpha and beta adaptin) and a ~50 and ~17 kDa subunit. There are two alpha-adaptin genes, alpha A and alpha C which have a tissue specific pattern of expression.
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