Use of spectrophotometer for ELISA
Posted 21 November 2009 - 10:15 PM
Thanks so much, anything you can help with would be great.
Posted 21 November 2009 - 11:22 PM
But then the microplate reader was born, and things changed a bit - made it quite a bit faster as well.
The only thing I'd be concerned about if I were you is the transfer process, and how that might make your results inaccurate. Obviously you're familiar with the ELISA principle - since the antibodies that are at the core of the ELISA reaction are bound to the microwell plate, I'm not sure how everything would respond if you transferred the solution and just left the antibodies there.
Sorry I'm not much help, I've just very little experience with this. But, you say that your mentor has done this before, so you should be fine as long as your mentor has done it with quantitative ELISA. I assume that you are looking for quantitative results?
Best of luck with science fair, by the way. ELISA is quite advanced for a student, so I congratulate you. I hope others can help you.
Edited by prof. moriarty, 21 November 2009 - 11:24 PM.
Posted 22 November 2009 - 03:50 AM
you will dilute ca 250 ul to 2000 ul, a 8 times dilution but the ligtpath will increase from ca 3mm to 10 mm thus the overall dilution will be about a factor 2.5.
This will influens your sensivity of the assay a little.
I would make the dilutions with the substate buffer but water also will work.
Edited by Gerard, 22 November 2009 - 03:53 AM.
Pluralitas non est ponenda sine necessitate
-- "You must assume no plural without necessity".
Posted 23 November 2009 - 11:52 AM
genius does what it must
i do what i get paid to do