chambered cell culture-slide for confocal microscopy - (Sep/15/2011 )
I am growing cells for imaging under confocal microscopy, previously I have been growing the cells on circular cover slips in 24 well plates, stimulating each well under different conditions, then staining for my protein of interest. (To preserve antibody I use staining technique in which I place a 20ul drop of antibody on some parafilm, remove the coverslip from the well, and place cell-side down on to the drop of antibody, the drop spreads to coat each cell for 15 mins and I use far less antibody than if I added it directly to the well.) However this results in very fiddly technique trying to get each slide out of the well, then back and forth between well and antibody for the washes.
Recently someone else in my lab suggested I use culture-slides (as in linkbelow), where I buy glass slides with plastic 8-well chambers glued on top, then I can culture the cells in these wells, stain induvidually, then remove the plastic chambers and affix a glass coverslip over the top of the cells.
Does anyone have any experience with using these culture-slides? especially with confocal microscopy where the thickness of samples can be very important? I am interested in making my working life much easier and less time consuming, but only if it still results in reproducable results
Yes, they are quite good. Very expensive though. The chambers are separated by a hydrophobic strip that should repel solutions if you remove the chamber before staining. I used to remove the chambers without the methanol step, and it worked fine.
Incidentally, i have found that if you use a hypodermic needle (gauge 18 works well), to lift the edge of a coverslip in a well, and then a pair of forceps to hold the coverslip, it is much easier than just using forceps.
The hydrophobic strip seperating the wells can become leaky, resulting in mixing of your antibodies. Particularly during overnight incubations. Better use the same primary for all wells. Also the capillay effect is more pronounced in the squared wells. You'll see rather heterogeneous distribution of your cells. Apart from that, sample thickness has never been an issue for me.
hmm I have also mentioned this to someone in the lab who used this previously, and they also mentioned that they couldn't get their samples into focus when they looked at them under confocal at first, and they had to scrape off any residual glue from the removal of the wells in order to get the cover slip close enough to the cells to be able to see them in focus.
Thanks for the advice though, I havent looked up the price yet (hate all these companies who force you to ring them up to get prices) but if they are that much more expensive then maybe it wouldn't be as worth it. I will look into it some more