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Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Mouse Semaphorin 7a.
(Peptide available as ab30844.)
Our Abpromise guarantee covers the use of ab23578 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||Use a concentration of 1 µg/ml. Detects a band of approximately 50,75,100 kDa (predicted molecular weight: 75 kDa).|
|ICC/IF||Use a concentration of 1 - 25 µg/ml.|
|IHC-FoFr||1/100. (PMID 19741144)|
|IHC-Fr||1/100. (PMID 19741144)|
|IHC-P||1/100. (see Abreview)|
|IP||Use at an assay dependent concentration.|
ab23578 detecting Semaphorin 7a in developing mouse neurons, differentiated from neural precursor cells cultured from embryonic day 13 neocortex; neurons were cultured on poly-D-lysine in the presence of Neurobasal Medium containing B27 supplement. Immunocytochemistry: all steps were performed in PBS. Cultures were fixed in 4% PFA for 15min, permeabilised with 0.1% Triton X-100 for 10min and blocked with 5% BSA, 0.1% Triton X-100 for 45min. ab23578 was incubated at 5µg/ml (12h in 5% BSA, 0.1% Triton X-100) at 4°C. Goat anti-rabbit AlexaFluor 488 was used as secondary antibody at 1/400 for 1hr at RT (in 5% BSA, 0.1% Triton X-100). Treated cultures were mounted on glass coverslips with Mowiol. ab23578 immunostaining (green colour) was neuronal and for the most part at the membrane with some cytosolic staining observed. No nuclear staining was observed, confirmed with a nuclear counterstain (To-pro-3; data not shown).
ab23578 detects Semaphorin 7a protein at 75kDa by WB. Semaphorin 7a is a protein that is tethered to the cell membrane via a GPI-anchor and is also glycosylated, therefore it is not surprising that we see some additional bands at ~50 and 100 kDa which are likely to be cleavage fragments and glycosylated protein respectively.