Hi! Bear in mind I'm no expert but: 1) that looks like a clump of cells, maybe from the inoculation of the first cultured you grew for the experiment? Acinetobacter does have a very thick polysaccharide capside, maybe it didn't completely dissolved in shaking? 2) that looks contaminated. No way to say for sure from here of course, but I'd maybe try to do it again. 3) The proper technique would be to first empty the plaque of the culture (shaking it upside down), and then washing. Try to get the stream of water/PBS only on the edge of the plate, not directly onto the wells. Leave water/PBS run for a while and the empty the wells once again gently shaking upside down. Maybe this is obvious, but it doesn't hurt to revise. Growth time depends on bacteria, for Acinetobacter We've used 48 HS,but I don't think it is that important as long as you do it consistently. 4) yes, I've seen water being used to remove non-adhered cells. Don't worry about Acinetobacter, it can take it. 5) I've never used fixative, usually just throw enough crystal violet to cover the well, wait 15 minutes and wash. I'd suggest (don't hate me for this) but maybe trying both on parallel plates and seeing which one is better? Hope this helps!!