Acridine orange adventures

[Wes]

Acridine orange is a fluorescent dye that can be excited with blue-cyan light and upon binding do DNA and RNA it fluoresces in green and red respectively (there are of course other things that make it glow green or red). Here I added the tiniest drop (< 100 nanoliters) of an acridine orange stock solution to a drop of pond water and captured a few images I decided to share. All images were acquired with the following fluorescent setup: 450-490 bandpass excitation filter, 510 dichroic mirror and 515 longpass filter.


A paramecium I think. Plan 16/0.32


Merismopedia. Neofluar 40/0.75


Colonial algae. Neofluar 40/0.75

[75RR]

wonderful images ... well done.

I see the upgrade you mentioned is paying dividends!

viewtopic.php?f=24&p=73559#p73559

[Wes]

Thanks 75RR. Its very refreshing to learn new techniques.

[PeteM]

Very cool images.

And seems even micro-holiday spirited: deck the slides with drops of acridine, fa la la la la la . . .?

[Wes]

Thanks Pete. The green and red touch certainly adds to the Christmas spirit.

[charlie g]

Thanks for these beautiful results, Wes. Was that first image a ciliate, and was that protozoan still active ( as in a gentle vital strain treatment with Janus Green)?

It would be good to know how long you exposed these pond organisms to your dilute stain ( approx. that is), before these stain effects manifested. Thanks for sharing this bench work, Wes.

Charlie guevara

[Wes]

Thanks Charlie

The majority of ciliates I spotted were moving around actively. The samples were exposed to AO from anywhere between 10 to 40 minutes. I think the intense illumination is what kills or at least impairs them after a period of observation. A rather dramatic end meets amoeba species stained with AO. They possess intensely fluorescing lysosomes (stained orange due to the low pH) and after about a minute or less of observation the lysosomes suddenly dissolve and the cell disintegrates in the matter of seconds. Below is an example before it disappeared.

[billbillt]

BEAUTIFUL IMAGES!...

THE BEST,
BILLT

[tgss]

Awesome shots Wes!
tgss

[Wes]

Thank you billt and tgss!