Q1) in principle I will assume that will be more than enough. However, it will of course depend on your cells and how "efficient" they are in exosome production. it is also important to pay attention to the exosome harvest and collect the exosomes at the right time. which time point is correct will depend on the cell type. One way of checking this is actually to use e.g. Dynabeads CD9 and pull out a small fraction of exosomes each day (without any pre-enrichment) during cell culture and stain for CD9 and perform analysis by flow cyt. this will give you an idea of exosome release and which would be the best period of performing exosome harvest and produce exosomes suitable for negative stain, immuno labelling and TEM analysis.
Q2) If you are not able to detect exosomes using CD9 this could be explained either by too little starting material, exosomes not expressing CD9 or the use of reducing conditions during sample prep for WB - as we have tested this WB clone for several years on many different samples and know that it works very well on exosomes. would have to look at your protocol in more detail to see if there are something to change. you may send it to me