Pan RAB (Internal Region) antibody

Details for Product No. ABIN572061, Supplier: Log in to see
Antigen
Epitope
Internal Region
Reactivity
Human, Mouse (Murine), Rat (Rattus)
Host
Rabbit
Clonality
Polyclonal
Application
Immunohistochemistry (IHC), Western Blotting (WB)
Options
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'Independent Validation' Badge
Antigen RAB
Lot Number Rb1082-020809-WS
Method validated Immunocytochemistry
Positive Control HeLa
Negative Control no primary or no secondary antibody
Notes ABIN572061 was successfully validated for ICC based the staining pattern consistent with published data.
Primary Antibody ABIN572061
Secondary Antibody goat anti-rabbit IgG (H+L) secondary antibody, Alexa Fluor 546 (ThermoFisher, A-11035)
Protocol
  • Grow HeLa cells in DMEM growth medium at 37°C in 5% CO2 to 70% confluency.
  • Fix cells in warm 4% formaldehyde for 20min at RT.
  • Wash cells twice with cold PBS.
  • Block and permeabilize cells in B-PBST (PBS, 1% BSA, 0.25% Triton X-100, 0.22µm filter sterilized) for 30min at RT.
  • Incubation with primary antibody ABIN572061 (antibodies-online, lot Rb1082-020809-WS) diluted 1:50 and 1:200 in B-PBST for 1h at RT.
  • Wash cells twice with PBS for 2min at RT.
  • Incubation with goat anti-rabbit IgG (H+L) secondary antibody, Alexa Fluor 546 (ThermoFisher, A-11035) diluted 1:500 in B-PBS for 1h art RT in the dark.
  • Wash cells thrice for 2min at RT in the dark.
  • Mount in VECTASHIELD HardSet Antifade Mounting Medium with DAPI (Vector laboratories, H-1500, lot ZC0816).
  • Confocal imaging on Zeiss LSM 880 at 63x magnification.
    Experimental Notes
    • ABIN572061 works very well in HeLa cells at both dilutions tested, 1:50 & 1:200. The strength of the signal at the 1:200 dilution suggests that it can be diluted even higher to 1:500 or 1:1000 without major loss in image quality.
    • A rabbit anti-giantin antibody in combination with the same secondary antibody as described in the protocol showed the expected localization. Also, both negative controls – no primary or no secondary antibody control – did not show any staining, indicating that the staining pattern shown in the image was indeed due to specific binding of ABIN572061 to its antigen(s).
    Validation Images
    Immunocytochemistry validation image for anti-Pan RAB (Internal Region) antibody (ABIN572061) Staining of HeLa cells with ABIN572061 in conjunction with an AF546 conjugated goat a...
    Immunogen A synthetic peptide from internal region of mouse Ras-related protein Rab-3A (RAB3A) conjugated to an immunogenic carrier protein has been used as the antigen. The antigen is homologous or shares a significant identity with many RAB molecules including RAB1, RAB2, RAB4, RAB9, RAB10, RAB11, RAB14, RAB26, RAB30, RAB35, RAB39, RAB43.
    Specificity Specific for RAB.
    Purification Whole serum
    Background Function: Involved in exocytosis by regulating a late step in synaptic vesicle fusion. Could play a role in neurotransmitter release by regulating membrane flow in the nerve terminal.
    Subcellular location: Cell membrane, Lipid-anchor, Cytoplasmic side Also known as: RAB3, RAB4, RAB9, RAB43, RAB35, RAB39, RAB30, RAB1, RAB4, RAB14, RAB11, RAB10, RAB26, RAB2.
    Application Notes Use at a dilution of 1:300 to 1: 2000.
    The optimal dilution should be determined by the end user.
    Not yet tested in other applications.
    Restrictions For Research Use only
    Format Lyophilized
    Reconstitution Reconstitute in 100 µL of sterile water. Centrifuge to remove any insoluble material.
    Handling Advice Avoid freeze and thaw cycles.
    Storage 4 °C/-20 °C
    Storage Comment Maintain the lyophilised/reconstituted antibodies frozen at -20°C for long term storage and refrigerated at 2-8°C for a shorter term. When reconstituting, glycerol (1:1) may be added for an additional stability. Avoid freeze and thaw cycles.
    Expiry Date 12 months
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