Good day to the forum members. Please I need help with my experiment on the worms. Am trying to make 2% agar pads that would be used for mounting the worms. I have gotten the protocol from wormbook.org and it was mentioned there that 5% agar solution was used, except that the name of the agar used is not mentioned. Please can anyone help me out with this agar to use. I am trying to carryout an ROS production test on the worms. Thank you in anticipation.
just to be clear, are you perhaps reading different sections/protocols where people describe 5% agar pads and 2% agarose pads? In any case, the agar we use works fine for pads (Agar Agar Kobe I)
OK,
Okayforum asked via the Moderator the following:
‘…Yes I plan to used the 2% agar pad made from 2%agar solution method. You mentioned 2% agarose pad. Is that different from the 2% agar pad I plan to use?’
So, to the first question, the answer is chemically; yes. Also the physical attributes of agarose are different, but I guess that’s not what you are getting at right?
The point is that there are a swath of different methods. A short summary of which would include:
2% agar (apparently used by someone at Cologne Uni according to this posting: http://www.researchgate.net/post/How_does_one_mount_caenorhabditis_elegans_on_a_slide_and_is_there_any_simple_stain_to_observe_them
3% agar pads - Shakes et al. and others for immunofluorescence.
4% agar pads: http://www.wormbook.org/chapters/www_intromethodscellbiology/intromethodscellbiology.html#d0e2311 (Protocol 28).
5% agar pads (same reference, Protocol 1)
And of course, 2% agarose pads as previously mentioned. I would try them out and see what suits your set up.
Steve
Thank you so much once again moderator. I was getting my self confused previously but now as regards the ROS production test I intend to carryout on the worms, since I would want the worms to be alive, just static while I take the image capture, I want to use the 5% agar solution method described in wormbook.org. Just to be completely clear, the agar I am supposed to use for this metho
d is Agar Agar Kobe I right?
Science is all about experimentation. If you’ve got some agar and agarose already in the lab, why spend more money when you can just do a little experiment yourself and find what condition works best for you.
I do intend to continue experimenting Snug, but then I need to confirm the type of agar i want to use.
just try using (as Snug said) some agar (that’s still in date) out of / on the chemicals cupboard/shelf and see how it goes…
remember, how static the worms will be depends on whether you anesthetise them…enough…
be sure to not use sodium azide, too
Alright, thank you very much for the vital information provided to me.