Hi guys,

I am trying to produce RNS with the guidance of this protocol :

To produce ONOO− , 10 mL NaNO2 (50 μmol/mL) and 10 mL H2O2 (25 μmol/mL) were mixed and stirred for 3 min. Then, 5 mL HCl (1 mol/L) and 5 mL NaOH (1.5 mol/L) were added the solution within 1 s. The mixed solution was stirred continuously until the solution changed to light yellow. The whole steps were carried out in an ice bath. ONOO− has strong absorbance at 302 nm. (https://doi.org/10.1002/adma.202206779)

Unfortunately, I can not make my solution into yellow color. I think there must be something tiny that I ignored caused this failure.

Is there anyone that can share this experiment details with me? or is there any other way to produce RNS in vitro?

Thank you.