The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Use a concentration of 1 µg/ml. Detects a band of approximately 56 kDa (predicted molecular weight: 44 kDa).
Use a concentration of 5 µg/ml.
Plays an important role in caveolae formation and organization. Required for the sequestration of mobile caveolin into immobile caveolae. Termination of transcription by RNA polymerase I involves pausing of transcription by TTF1, and the dissociation of the transcription complex, releasing pre-rRNA and RNA polymerase I from the template. PTRF is required for dissociation of the ternary transcription complex.
Congenital generalized lipodystrophy 4
Belongs to the PTRF/SDPR family.
Phosphorylated. Present in active and inactive forms. Changes in phosphorylation pattern may alter activity. Five truncated forms are found in the caveolae. These are thought to be the result of proteolysis and may be phosphorylation-dependent.
Membrane > caveola. Cell membrane. Microsome. Endoplasmic reticulum. Cytoplasm > cytosol. Mitochondrion. Nucleus. Found at the surface of the caveolae. Also found in the plasma membrane, microsomal and cytosolic fractions and at a low level in the mitochondrial and nuclear fractions. Translocates to the cytoplasm from the caveolae upon insulin stimulation. CAV1 is necessary to recruit it to the cell membrane (By similarity). Co-localizes with CAV1 in lipid rafts in adipocytes.
PTRF contains a number of potential phosphorylation sites (SwissProt data) which may explain it running at a higher molecular weight than predicted. The 56 kDa band is comparable to molecular weights seen with other commercially available antibodies to PTRF.
ICC/IF image of ab78553 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 (ab78553, 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.
IHC image of PTRF staining in normal human liver formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab78553, 5ug/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.