The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml. Detects a band of approximately 16, 80 kDa (predicted molecular weight: 12 kDa).Can be blocked with Human Sumo 1 peptide (ab49766).
Use a concentration of 5 µg/ml.
Ubiquitin-like protein that can be covalently attached to proteins as a monomer or a lysine-linked polymer. Covalent attachment via an isopeptide bond to its substrates requires prior activation by the E1 complex SAE1-SAE2 and linkage to the E2 enzyme UBE2I, and can be promoted by E3 ligases such as PIAS1-4, RANBP2 or CBX4. This post-translational modification on lysine residues of proteins plays a crucial role in a number of cellular processes such as nuclear transport, DNA replication and repair, mitosis and signal transduction. Involved for instance in targeting RANGAP1 to the nuclear pore complex protein RANBP2. Polymeric SUMO1 chains are also susceptible to polyubiquitination which functions as a signal for proteasomal degradation of modified proteins. May also regulate a network of genes involved in palate development.
Involvement in disease
Defects in SUMO1 are the cause of non-syndromic orofacial cleft type 10 (OFC10) [MIM:613705]; also called non-syndromic cleft lip with or without cleft palate 10. OFC10 is a birth defect consisting of cleft lips with or without cleft palate. Cleft lips are associated with cleft palate in two-third of cases. A cleft lip can occur on one or both sides and range in severity from a simple notch in the upper lip to a complete opening in the lip extending into the floor of the nostril and involving the upper gum. Note=A chromosomal aberation involving SUMO1 is the cause of OFC10. Translocation t(2;8)(q33.1;q24.3). The breakpoint occurred in the SUMO1 gene and resulted in haploinsufficiency confirmed by protein assays.
Belongs to the ubiquitin family. SUMO subfamily. Contains 1 ubiquitin-like domain.
Cleavage of precursor form by SENP1 or SENP2 is necessary for function. Polymeric SUMO1 chains undergo polyubiquitination by RNF4.
Nucleus membrane. Nucleus speckle. Cytoplasm. Recruited by BCL11A into the nuclear body.
Lane 1: Wild-type HAP1 cell lysate (20 µg) Lane 2: Sumo 1 knockout HAP1 cell lysate (20 µg) Lane 3: HeLa cell lysate (20 µg) Lane 4: MCF-7 cell lysate (20 µg) Lanes 1 - 4: Merged signal (red and green). Green - ab49767 observed at 16 kDa. Red - loading control, ab8245, observed at 37kDa.
ab49767 was shown to recognize Sumo 1 when Sumo 1 knockout samples were used, along with additional cross-reactive bands. Wild-type and Sumo 1 knockout samples were subjected to SDS-PAGE. ab49767 at a concentration of 1 μg/ml and ab8245 (loading control to GAPDH) at a dilution of 1/10000 were incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.
Western blot - Anti-Sumo 1 antibody (ab49767)
Anti-Sumo 1 antibody (ab49767) at 1 µg/ml + A431 (Human epithelial carcinoma cell line) Whole Cell Lysate at 10 µg
ICC/IF image of ab49767 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab49767, 5µg/ml) overnight at +4°C. 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) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 4% PFA fixed (10 min) Hek293, HepG2 and MCF7 cells at 5µg/ml, and in 100% methanol fixed (5 min) HeLa, Hek293, HepG2 and MCF7 cells at 5µg/ml.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Sumo 1 antibody (ab49767)This image is courtesy of an Abreview submitted by Antibody Solutions Ltd.
ab49767 staining Sumo 1 in Human placenta tissue sections by IHC-P (formaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 5 minutes of peroxidase block followed by 10 minutes of protein block at 20°C; antigen retrieval was heat mediated in retrieval solution. Samples were incubated with primary antibody (1/250 in antibody diluent) for 45 minutes at 20°C. An undiluted HRP-conjugated polymer goat anti-mouse/rabbit IgG polyclonal polymer was used as the secondary antibody.