Secretory protein? - How to determine if my protein is surely secreted? (Jul/28/2009 )

Dear all:

Recently, I have been investigating a novel protein in the fly Drosophila.
Based on following observations, I speculated it can be secreted by epithelium cells.

1. My protein is predicted to contain a signal peptide at the N-terminal.

2. Using immunostaining, this protein can colocalized with Rab7 (a late endosome marker) within cells that could be endocytosing it.

However, due to a lack of antibodies for secretory vesicles, I just wonder if I can use GFP-tagged Golgi marker instead.

If this protein indeed colocalized with Golgi, can I say that it is been secreting?

Moreover, what other experiments should I do in the future, in order to convince other that this protein is indeed been secreted?

Thanks in advance for all your advice.

Have you tried WB with concentrated and dialysed samples from the culture medium?

genehunter on Jul 29 2009, 12:27 AM said:

Yeah, I had a plamid containing C-termianl V5 tagged version of that protein.
I will try what you say in the future.