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Reusing the black 96-well plate for reading luminescence - For luciferease reporter assay (Jul/20/2010 )

Dear all,

I am wondering if any of you have experience in reusing the black/white 96-well plate for luminescence reading. Someone told me once before that i could wash the plates and reuse them...is it true? Suppose what i need to do then is to make sure i have washed everything out thoroughly with detergent?


I could have use brand new plates for each and every assay, if my lab is rich enough...


Thanks!

-WYF-

WYF on Jul 20 2010, 08:05 AM said:

Dear all,

I am wondering if any of you have experience in reusing the black/white 96-well plate for luminescence reading. Someone told me once before that i could wash the plates and reuse them...is it true? Suppose what i need to do then is to make sure i have washed everything out thoroughly with detergent?


I could have use brand new plates for each and every assay, if my lab is rich enough...


Thanks!


Ehh, I think it depends on how sensitive your work is, and the signals you're getting compared to background. Try running a new plate, then thoroughly washing it and re-running the same experiment on the washed plate after it has dried. I would not recommend using a new plate twice (or more) since proteins that have adhered to the plate walls may have a tendency to stick on pretty tough.

-PipetteTip-

Thanks!



Ehh, I think it depends on how sensitive your work is, and the signals you're getting compared to background. Try running a new plate, then thoroughly washing it and re-running the same experiment on the washed plate after it has dried. I would not recommend using a new plate twice (or more) since proteins that have adhered to the plate walls may have a tendency to stick on pretty tough.



Thanks! I agree that the only way to find out is to test it myself. I am just slightly concern with the cost. The luciferase reagent is so expensive! That's why i am hoping to get some advice from experts through this forum...

-WYF-

precisily then why u shud think twice before reusing the plate!!! one is the cost of the reagent and the another thing is how much more difficult it is going to make ur trouble shooting in case sumthing goes wrong with the assay!!!!
having said tat io also agree with PT tat if the values are way too high abouve the background of the plate then u can reuse it.. how many number of times tat depends on ur samples...

Another alternative is run the blank plate before loading the samples and then check the background signal each time for all wells... and then u will know how robust it is!!!!

-Prep!-

Prep! on Jul 22 2010, 04:53 AM said:

precisily then why u shud think twice before reusing the plate!!! one is the cost of the reagent and the another thing is how much more difficult it is going to make ur trouble shooting in case sumthing goes wrong with the assay!!!!
having said tat io also agree with PT tat if the values are way too high abouve the background of the plate then u can reuse it.. how many number of times tat depends on ur samples...

Another alternative is run the blank plate before loading the samples and then check the background signal each time for all wells... and then u will know how robust it is!!!!


Brilliant! Run a blank plate! Why haven't i thought about that! Thanks a bunch!

-WYF-

I'm used to recycle plates without problems.

A blank plate is a good idea but it is not enough: the problem in the sample contamination is not that it will still give luminescence (after 24-48 hrs luciferase is completely disrupted). In contrast, some remaining chemicals (i.e., the stop solution) could interfere with subsequent readings. I carefully wash the plates with a mild detergent (like SDS 0.1%) and flush them very well with distilled water. Just let them dry at RT as with higher temperatures (we have a 50 C incubator) the plastic could deform a little bit and you will have problems as the detector does not fit exactly the well.

Beware to run your readings at room temperature, in my blog I explained why.

-96well-