We have homogenates of dorsal and ventral hippocampi of male and female adolescent rats and need to determine NPY levels by ELISA (Ray-Biotech). We can not find specifications for sample dillution.
Hi Deborah. In brief I do not know the concentrations or dilutions you could try.
However I have ran a number of Elisa with samples that I do not know the target protein/molecule concentration.
In these instances I have run an Elisa "test strip".
Elisa are generally composed of 12 x 8 "well" strips. Before running the full Elisa I would suggest using one of these strips. On the have 3 points of the 8 point standard curve (i.e. S1, S5 and S8). The remaining five wells can contain a range of possible dilutions of your homogenates (i.e. neat, 1:10, 1:20 1:50 1:100).
Data from the test strip will then inform on the dilutions to be used for the remaining strips/wells.
I hope this helps (and that it makes sense!)
Gary
I would recommend you to do a curve of dilution, because the amount of protein varies a lot depending the amount o tissue you have extracted.
So, my suggestion is to run a protein assay and after that you can run a curve (1:1. 1:10, 1:50, :100 and so on) and see how is the signal. After that you can decide which concentration is the best for you.
let me know if you have any question, you can email me ([email protected])
cheers, Doug.
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