Anti-PADI4 / PAD4 antibody (ab38772)

Western blot - PADI4 / PAD4 antibody (ab38772)

All lanes : Anti-PADI4 / PAD4 antibody (ab38772) at 1/250 dilution

Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
Lane 2 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate (ab7899)
Lane 3 : A431 (Human epithelial carcinoma cell line) Whole Cell Lysate (ab7909)
Lane 4 : HEK293 Human embryonic kidney cell line Whole Cell Lysate (ab7902)
Lane 5 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate (ab7900)

Lysates/proteins at 10 µg per lane.

Secondary
IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution

Performed under reducing conditions.

Predicted band size : 74 kDa
Observed band size : 67 kDa (why is the actual band size different from the predicted?)

Full length PADI4 has been shown to migrate at 67 kDa.  Please refer to the reference below for further details.

Chang X et al. Localization of peptidylarginine deiminase 4 (PADI4) and citrullinated protein in synovial tissue of rheumatoid arthritis. Rheumatology (Oxford) 44:40-50 (2005). PMID:15466895.

Immunocytochemistry/ Immunofluorescence - PADI4 / PAD4 antibody (ab38772)

ICC/IF image of ab38772 stained human HeLa cells. The cells were PFA fixed (10 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab38772, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - PADI4 / PAD4 antibody (ab38772)

ab38772 staining PADI4 / PAD4 in human spleen tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with 10% NBF and a heat mediated antigen retrieval step was performed using TRIS/EDTA pH 9.0. Samples were then blocked with 0.25% casein in PBS for 5 minutes at 25ºC followed by incubation with the primary antibody at a 1/400 dilution for 30 minutes at 25ºC (serum free antibody diluent). An undiluted HRP-conjugated goat polyclonal was used as secondary antibody.

Image courtesy of Ross Benik by Abreview.