The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
ICC/IF: Use at a concentration of 1 µg/ml.
IHC-FoFr: 1/300 (See abreview for more details)
WB: Use at a concentration of 1 µg/ml. Detects a band of approximately 60,45 kDa (predicted molecular weight: 60,58,48 kDa).
Not yet tested in other applications.
Optimal dilutions/concentrations should be determined by the end user.
May function as a growth and differentiation factor involved in neuritogenesis. May induce ERBB3 activation.
Restricted to brain (at protein level).
Contains 1 EGF-like domain.
Expressed in brain of 3 months, 5 and 10-year-old individuals.
N-glycosylated. O-glycosylated; contains chondroitin sulfate glycans. Part-time proteoglycan, expressed in part as a proteoglycan exhibiting chondroitin sulfate glycans and in part as a non-proteoglycan form. The relative amount of both forms depends on tissues and tissues maturation. Phosphorylated; in intracellular and extracellular parts.
Cell membrane. Endoplasmic reticulum membrane. Golgi apparatus membrane. In neurons, localizes to synaptic junctions (By similarity). Also detected in the endoplasmic reticulum and the Golgi (By similarity). Partially enriched in lipid rafts.
It is impossible to conclude from the data whether the higher observed band is Neuroglycan C isoform 3 (60 kDa) or Neuroglycan C isoform 1 (57 kDa). The epitope for this antibody is present in all 3 isoforms of the Neuroglycan C protein. We suspect the lower band is Isoform 2.
Western blot - Neuroglycan C antibody (ab31946)
All lanes : Anti-Neuroglycan C antibody (ab31946) at 1 µg/ml
Lane 1 : Brain (Mouse) Tissue Lysate Lane 2 : Brain (Rat) Tissue Lysate - normal tissue
Lysates/proteins at 10 µg per lane.
Secondary All lanes : IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution
Immunocytochemistry/ Immunofluorescence - Neuroglycan C antibody (ab31946)
ICC/IF image of ab31946 stained human HeLa cells. The cells were methanol fixed (5 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab31946, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. 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). DAPI was used to stain the cell nuclei (blue).
ab31946 staining Neuroglycan C in rat brain tissue section by Immunohistochemistry (PFA perfusion fixed frozen sections). Tissue from 4% PFA perfused animals underwent overnight fixation in 4% paraformaldehyde, cryoprotected in 30% sucrose and cut using cryostat.The primary antibody was diluted, 1/300 (PBS + 0.3% Triton X100) and incubated with sample for 18 hours at 20°C. An Alexa Fluor® 488 conjugated goat polyclonal to rabbit IgG was used at 1/1000 dilution, as secondary.
This image is courtesy of an Abreview submitted by Dr Sophie Pezet.