sodium azide in ELISA

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Hatuey
Hatuey's picture
sodium azide in ELISA

Hello,
I need to run an ELISA and measure a control solution that contains sodium azide. Sodium azide inhibits completely HRP. Some ELISA kits contain a component in the solution of the HRP conjugate that prevents inhibition of HRP by sodium azide. Does any one knows what component(s) is that?

Sami Tuomivaara
Sami Tuomivaara's picture
Hatuey,

Hatuey,

Could you please let us know what kits have components that prevents HRP inhibition by azide, I haven't heard of this!

I think that azide in your sample (or control), blocking buffer or antibody buffer doesn't matter as long as your washing buffer doesn't have azide in it. The final washing before adding substrate should get rid of any small molecules such as azide.

Cheers,
Suola

Dr._Eldon_Tyrell
Dr._Eldon_Tyrell's picture
 It's also the first time I'm

 It's also the first time I'm hearing of that to be a problem..
As Suola says, why does this matter to you? If, for example, your sample contains SA you would wash it away before staining.

And what concentrations are we talking about anyway? 

As long as you have no SA in your HRP-Antibody storage buffer it should not be a problem..

Valérie
Valérie's picture
Hi, I am developing an ELISA

Hi, I am developing an ELISA for which I have to use 2%goat sera diluted in PBS 1X ; 0.1% tween 20 to avoid non specific binding.
I have tested 2 different goat sera, one with sodium azide (0.08%) the other one without. 
With the one that not contain sodium azide my background is too high.
My background is acceptable (<0.1) with a 2%goat sera that contain 0.08% sodium azide + PBS1X;0.1%tween20 as preservative agent.
I have try to dilute my HRP antibody only in (PBS 1X, tween 0.1%) but in that case I have some insconsistent OD.
It seems that if I want to avoid goat sera screening (It may takes time and I am under rush) I have to know If 0.08% of sodium azide is even to high to dilute my HRP with.
Note that my goat sera is used at a final 2% my 0.08 is only 0.0016% no?
Please let my know if you have anything about that

bobhardcastle
bobhardcastle's picture
If you are using a hetrogeous

If you are using a hetrogeous assay (ie includes wash steps between antibody incubations) then azide in any buffer except your HRP dilution buffer should not make a big impact on your assay signal.

What is the intended purpose of the inclusion of azide in your buffer, as a preservative?

If this is the case you could try using a different preservative that are known not to inhibit HRP, such as Kathon or thimerosal.