Immunocytochemistry/ Immunofluorescence - Clathrin antibody [X22] - Membrane Vesicle Marker (ab2731)

ab2731 staining human HEK 293 cells by ICC/IF.  Cells were PFA fixed and permeabilized in 0.01% Triton X-100 in PBS prior to blocking in 1% BSA for 1 hour at 22°C.  The primary antibody was diluted 1/75 and incubated with the sample for 1 hour at 22°C.  An Alexa Fluor® 405 conjugated goat anti-mouse antibody diluted 1/400 was used as the secondary.

This image is courtesy of an Abreview submitted by Mrs Danijela Markovic

Western blot - Clathrin antibody [X22] - Membrane Vesicle Marker (ab2731)

All lanes : Anti-Clathrin antibody [X22] - Membrane Vesicle Marker (ab2731) at 1/500 dilution

Lane 1 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate
Lane 2 : Raji (Human Burkitt's lymphoma cell line) Whole Cell Lysate

Lysates/proteins at 10 µg per lane.

Secondary
Goat polyclonal to Mouse IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

Observed band size : 180 kDa (why is the actual band size different from the predicted?)
Additional bands at : 240 kDa,450 kDa. We are unsure as to the identity of these extra bands.

Immunocytochemistry/ Immunofluorescence - Clathrin antibody [X22] - Membrane Vesicle Marker (ab2731)

ab2731 staining Clathrin - Membrane Vesicle Marker in Rat hypothalamus primary cells by ICC/IF (immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.5% TX-100 and blocked with 5% serum for 20 minutes at 25ºC. Samples were incubated with primary antibody (1/500 in 1% goat serum; 0.1% TX100; PBS) for 16 hours at 4ºC. An Alexa Fluor^®546-conjugated Goat polyclonal to mouse IgG, dilution 1/500, was used as secondary antibody.

This image is courtesy of an Abreview submitted by Dr Vladimir Milenkovic

Immunocytochemistry/ Immunofluorescence - Clathrin antibody [X22] - Membrane Vesicle Marker (ab2731)

ab2731 staining Clathrin in pig retinal pigment epithelium (RPE) cells by Immunocytochemistry/ Immunofluorescence. Cells were fixed with paraformaldehyde, permeabilized with 0.5% Triton ×100 and blocked with 5% serum for 20 minutes at 25⁰C. Samples were incubated with primary antibody (1/500: in 0.1% TX100, 1% goat serum, 1X PBS) for 16 hours at 4°C. An Alexa Fluor^®546-conjugated goat polyclonal to mouse IgG was used as secondary antibody at 1/500 dilution.

This image is a courtesy of Vladimir Milenkovic

Flow Cytometry - Anti-Clathrin antibody [X22] - Membrane Vesicle Marker (ab2731)

ab2731 staining clathrin in Human platelets by Flow Cytometry. Platelets were isolated form Platelet rich plasma and suspended in PBS, fixed with paraformaldehyde and permeabilized with 0.1% Triton-X100 in 2% BSA for 30 minutes. The sample was incubated with the primary antibody (1/200 in PBS + 2% BSA) for 18 hours at 4ºC. An Alexa Fluor^®594-conjugated Goat anti-mouse IgG polyclonal (1/500) was used as the secondary antibody.
Gating Strategy: Platelets

This image is courtesy of an Abreview submitted by Mahesh Shivananjappa