The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 5 µg/ml.
1/20 - 1/50.
Use 1µg for 106 cells.
Involved in melanosome biogenesis by ensuring the stability of GPR143. Plays a vital role in the expression, stability, trafficking, and processing of melanocyte protein PMEL, which is critical to the formation of stage II melanosomes.
Expression is restricted to melanoma and melanocyte cell lines and retina.
Endoplasmic reticulum membrane. Golgi apparatus. Golgi apparatus > trans-Golgi network membrane. Melanosome. Also found in small vesicles and tubules dispersed over the entire cytoplasm. A small fraction of the protein is inserted into the membrane in an inverted orientation. Inversion of membrane topology results in the relocalization of the protein from a predominant Golgi/post-Golgi area to the endoplasmic reticulum. Melanoma cells expressing the protein with an inverted membrane topology are more effectively recognized by specific cytolytic T-lymphocytes than those expressing the protein in its native membrane orientation.
IHC image of ab785 staining in Human Skin Melanoma 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 ab785, 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.
Overlay histogram showing Malme-3 cells stained with ab785 (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. The cells were then incubated with the antibody (ab785, 1µg/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. Acquisition of >5,000 events was performed. This antibody gave a positive signal in Malme-3 cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.
Suzuki S et al. The Origin and Role of Autophagy in the Formation of Cytoplasmic Granules in Canine Lingual Granular Cell Tumors. Vet PatholN/A:N/A (2014).
Read more (PubMed: 25161210) »
Civenni G et al. Human CD271-Positive Melanoma Stem Cells Associated with Metastasis Establish Tumor Heterogeneity and Long-term Growth. Cancer Res71:3098-109 (2011).
Read more (PubMed: 21393506) »