The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration.
1/50000 - 1/200000. Detects a band of approximately 150 kDa (predicted molecular weight: 139 kDa).
1/100 - 1/250. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Application notesIs unsuitable for Flow Cyt or IP.
FunctionRequired for normal progression of S-phase. Involved in the circadian rhythm autoregulatory loop. Negatively regulates CLOCK-NPAS2/BMAL1-induced transactivation of PER1 possibly via translocation of PER1 into the nucleus. Promotes TIPIN nuclear localiZation. Involved in cell survival after DNA damage or replication stress. May be specifically required for the ATR-CHK1 pathway in the replication checkpoint induced by hydroxyurea or ultraviolet light. May also play an important role in epithelial cell morphogenesis and formation of branching tubules.
Tissue specificityExpressed in all tissues examined including brain, heart, lung, liver, skeletal muscle, kidney, placenta, pancreas, spleen, thymus and testis. Highest levels of expression in placenta, pancreas, thymus and testis.
Sequence similaritiesBelongs to the timeless family.
Immunocytochemistry/Immunofluorescence analysis of HT-29 (Human colorectal adenocarcinoma cell line) labeling Timeless with Purified ab109512 at 1/1000 dilution (5 µg/ml). Cells were fixed with 4% PFA and permeabilized with 0.1% tritonX-100. ab150077 Goat anti rabbit IgG(Alexa Fluor® 488) at 1/1000 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI. PBS was used instead of the primary antibody as the negative control.
Western blot - Timeless antibody [EPR5275] (ab109512)
All lanes : Anti-Timeless antibody [EPR5275] (ab109512) at 1/50000 dilution
Lane 1 : HeLa cell lysate Lane 2 : Jurkat cell lysate Lane 3 : JAR cell lysate Lane 4 : BxPC-3 cell lysate