Is there a best way to pick up cells from coony in cell line cuture from 6 wells plate ? I need to isolate some cells from colony /well from 6 plate wells and incubate them, freezing some other.
I put them on the microscope stage and pick them up with a 10 ul pipet tip while watching through the 10x lens. I know it doesn't sound like that would work, but once you try it, you'll be surprised how easy it is.
You can use paper cloning discs (Fisher Scientific catalog #07-907-10A). They come sterile and in different diameters (3.2, 4.8 & 6.4mm). Just remove the media from the wells and wash with PBS. Then soak the disc in trypsin and then place it (using a sterile forceps) over your colony/colonies for a few minutes. Pick the disc up (the cells will be stuck to the paper) and transfer it to a new well with media in it. Let it sit for a few minutes then shake the disc in the media to dislodge the cells.
Good Luck!
You can also try cloning rings. They are plastic or glass and you can easily clean them for reuse. They work the same way as paper cloning discs suggested by Tom. Since they are plastic or glass, they are heavier and easier to stand on your tissue culture dish. To prevent leaking of trypsin, you can use some sterile silicone grease (already included in Sigma cloning rings, cat# C1059-1EA, attached). Good luck!
http://www.sigmaaldrich.com/catalog/product/sigma/c1059?lang=en®ion=US
I put them on the microscope stage and pick them up with a 10 ul pipet tip while watching through the 10x lens. I know it doesn't sound like that would work, but once you try it, you'll be surprised how easy it is.
I have picked soft agar colonies using the way John described. That only works for colonies in suspension or in soft agar. I assuming the colonies Ahmed talked about were the ones attached to tissue culture plates.
Tom Masi, Ying Henderson, John Schloendorn, Thanks alot for these answers and for all this attention. Good luck for all .
This is very late, but in case anyone is still interested, here is a simple agarose overlay method:
https://www.youtube.com/watch?v=IhOP397sCC8
titled "cloning cells with the agarose method"
see the above youtube video; there is more information on my website (thelindberglab.com) or on the Addgene site. If you overlay your plate with melted agarose mixed with concentrated DMEM (no serum)(1x final), the cells will stick to the agarose and can be punched out using a sterile 200 ul blunt cut pipette (make your own and autoclave, or buy them). Just make sure the agarose solution is not too hot before pouring on plate- should be melted but not really hot.
just got back to Researchgate, sorry for late response. You should be able to use a sterile, cut-off 200 ul pipet tip to punch through the agarose using a pipettor. Dispense the punch into 200 ul rich medium in a 48 well plate and vigorously pipet up and down to shake the cells loose (they will also migrate out, if they prefer the plate). Feed the wells with more rich medium (supplement with an extra 10% extra serum if your cells need growth factors to get going). In my experience most cell lines will grow up within 2 weeks. While I have not worked much with cells in suspension I note that there is a similar method published for single cell S2 cells in the literature.
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