The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
ICC/IF: Use at a concentration of 1 µg/ml.
IHC-P: 1/500. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
WB: Use at a concentration of 1 µg/ml. Detects a band of approximately 35, 39 kDa representing two alternatively spliced forms of UNG that are differentially localized (predicted molecular weight: 33, 35 kDa).
Not yet tested in other applications.
Optimal dilutions/concentrations should be determined by the end user.
Excises uracil residues from the DNA which can arise as a result of misincorporation of dUMP residues by DNA polymerase or due to deamination of cytosine.
Isoform 1 is widely expressed with the highest expression in skeletal muscle, heart and testicles. Isoform 2 has the highest expression levels in tissues containing proliferating cells.
Involvement in disease
Defects in UNG are a cause of immunodeficiency with hyper-IgM type 5 syndrome (HIGM5) [MIM:608106]. Hyper-IgM syndrome is a condition characterized by normal or increased serum IgM concentrations associated with low or absent serum IgG, IgA, and IgE concentrations. HIGM5 is associated with profound impairment in immunoglobulin (Ig) class-switch recombination (CSR) at a DNA precleavage step.
Belongs to the uracil-DNA glycosylase family.
Isoform 1 is processed by cleavage of a transit peptide.
All lanes : Anti-UNG antibody (ab23926) at 1 µg/ml
Lane 1 : HeLa (Human epithelial carcinoma cell line) whole cell lysate Lane 2 :Jurkat whole cell lysate (ab7899) Lane 3 : HeLa (Human epithelial carcinoma cell line) nuclear lysate
Lysates/proteins at 20 µg per lane.
Secondary All lanes : Goat polyclonal to Rabbit IgG (Alexa Fluor® 680) at 1/10000 dilution
Predicted band size: 33, 35 kDa Observed band size: 35,39 kDa (why is the actual band size different from the predicted?) Additional bands at: 18 kDa (possible cleavage fragment), 18 kDa (possible cross reactivity), 23 kDa (possible cleavage fragment), 23 kDa (possible cross reactivity), 40 kDa (possible cross reactivity)
UNG is present in the cell in two isoforms, that differ at their amino-terminal ~40 amino acids. This results in one isoform that is present in the nucleus and one that is present in the mitochondria. This antibody strongly recognizes a doublet at approximately 35 and 39 kDa which closely correspond in size to these two isoforms (whose predicted molecular weights are 33 and 35 kDa).
ICC/IF image of ab23926stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab23926, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).
ab23926 staining UNG in human heart muscle. Paraffin embedded human heart muscle tissue was incubated with ab23926 (1/500 dilution) for 30 mins at room temperature. Antigen retrieval was performed by heat induction in citrate buffer pH 6.
ab23926 was tested in a tissue microarray (TMA) containing a wide range of normal and cancer tissues as well as a cell microarray consisting of a range of commonly used, well characterised human cell lines.
Su MT et al. Uracil DNA glycosylase BKRF3 contributes to Epstein-Barr virus DNA replication through physical interactions with proteins in viral DNA replication complex. J Virol88:8883-99 (2014).
Read more (PubMed: 24872582) »
Kitamura K et al. Uracil DNA glycosylase counteracts APOBEC3G-induced hypermutation of hepatitis B viral genomes: excision repair of covalently closed circular DNA. PLoS Pathog9:e1003361 (2013).
Read more (PubMed: 23696735) »