The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use 2µg for 106 cells. ab170190-Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
Use at an assay dependent concentration.
1/100. Predicted molecular weight: 273 kDa.
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Central component of the spliceosome, which may play a role in aligning the pre-mRNA 5'- and 3'-exons for ligation. Interacts with U5 snRNA, and with pre-mRNA 5'-splice sites in B spliceosomes and 3'-splice sites in C spliceosomes.
Involvement in disease
Defects in PRPF8 are the cause of retinitis pigmentosa type 13 (RP13) [MIM:600059]. RP leads to degeneration of retinal photoreceptor cells. Patients typically have night vision blindness and loss of midperipheral visual field. As their condition progresses, they lose their far peripheral visual field and eventually central vision as well. RP13 inheritance is autosomal dominant.
Contains 1 MPN (JAB/Mov34) domain.
The MPN domain has structural similarity with viral ribonucleases and RNase H, but unlike RNases, it does not bind any metal ions.
Phosphorylated upon DNA damage, probably by ATM or ATR.
IHC image of PRPF8 staining in Human Normal Cervix formalin fixed paraffin embedded tissue section, performed on a Leica Bond™ 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 ab51366, 5µg/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.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Western blot - PRPF8 antibody [2834C1a] (ab51366)
Anti-PRPF8 antibody [2834C1a] (ab51366) at 1/100 dilution + HeLa whole cell lysate at 25 µg
Secondary Anti mouse IgG antibody at 1/2500 dilution
Overlay histogram showing HeLa cells stained with ab51366 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab51366, 2µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Unlabelled sample (blue line). Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.