SpecificityWe have data to indicate that this antibody may not cross react with Xenopus laevis. However, this has not been conclusively tested and expression levels may vary in certain cell lines/tissues.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
1/10000 - 1/50000. Detects a band of approximately 75 kDa.
Use at 2-5 µg/mg of lysate.
Use a concentration of 5 µg/ml.
FunctionBinds both single-stranded and double-stranded DNA and promotes ATP-independent annealing of complementary single-stranded DNAs and D-loop formation in superhelical double-stranded DNA. May play a role in maintenance of genomic integrity.
Involvement in diseaseNote=A chromosomal aberration involving FUS is found in a patient with malignant myxoid liposarcoma. Translocation t(12;16)(q13;p11) with DDIT3. Note=A chromosomal aberration involving FUS is a cause of acute myeloid leukemia (AML). Translocation t(16;21)(p11;q22) with ERG. Defects in FUS may be a cause of angiomatoid fibrous histiocytoma (AFH) [MIM:612160]. A distinct variant of malignant fibrous histiocytoma that typically occurs in children and adolescents and is manifest by nodular subcutaneous growth. Characteristic microscopic features include lobulated sheets of histiocyte-like cells intimately associated with areas of hemorrhage and cystic pseudovascular spaces, as well as a striking cuffing of inflammatory cells, mimicking a lymph node metastasis. Note=A chromosomal aberration involving FUS is found in a patient with angiomatoid fibrous histiocytoma. Translocation t(12;16)(q13;p11.2) with ATF1 generates a chimeric FUS/ATF1 protein. Defects in FUS are the cause of amyotrophic lateral sclerosis type 6 (ALS6) [MIM:608030]. ALS6 is a familial form of amyotrophic lateral sclerosis. ALS is a neurodegenerative disorder affecting upper motor neurons in the brain and lower motor neurons in the brain stem and spinal cord, resulting in fatal paralysis. Sensory abnormalities are absent. Death usually occurs within 2 to 5 years. The etiology of amyotrophic lateral sclerosis is likely to be multifactorial, involving both genetic and environmental factors. The disease is inherited in 5-10%.
Sequence similaritiesBelongs to the RRM TET family. Contains 1 RanBP2-type zinc finger. Contains 1 RRM (RNA recognition motif) domain.
Post-translational modificationsArg-216 and Arg-218 are dimethylated, probably to asymmetric dimethylarginine.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human ovarian carcinoma (left) and mouse teratoma (right) tissues labelling TLS/FUS with ab70381 at 1/5000 (0.2µg/ml) and 1/1000 (1µg/ml). Detection: DAB.
Western blot - TLS/FUS antibody (ab70381)
All lanes : Anti-TLS/FUS antibody (ab70381) at 0.05 µg/ml
Lane 1 : HeLa whole cell lysate at 10 µg Lane 2 : HeLa whole cell lysate at 50 µg
Detection of Human TLS/FUS by Immunoprecipitiaton. ab70381 used at 3µg/10cm plate for IP in lane 2, other antibodies used in lanes 1, 3, 4, 5 & 6. Control IgG used in lane 5. Lanes 1-5 - cells from one half of one 10cm plate of of HeLa whole cell lysate, and lanes 6 & 7 100µg of HeLa whole cell lysate used. ab70381 used at 0.02µg/ml in WB.
Detection: Chemiluminescence with an exposure time of 5 seconds. Band size 58kDa.
ab70381 (1µg/ml) staining TLS in human colon using an automated system (DAKO Autostainer Plus). Using this protocol there is cytoplasmic and nuclear staining. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 AR buffer citrate pH 6.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
ICC/IF image of ab70381 stained HeLa cells. The cells were 100% methanol fixed (5 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 (ab70381, 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.
Murakami T et al. ALS mutations in FUS cause neuronal dysfunction and death in Caenorhabditis elegans by a dominant gain-of-function mechanism. Hum Mol Genet21:1-9 (2012).
IHC (PFA fixed)
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