ROS-Glo H2O2

- 200 ul H2O2 Substrate, 10 mM

- 500 ul Signal Enhancer Solution
- 500 ul D-Cysteine, 100X
- 10 ul H2O2 Substrate Dilution Buffer
- 1 vial Luciferin Detection Reagent
- 50 ml Reconstitution Buffer

*Reconstituted Luciferin Detection Reagent (RLDL) = Reconstitution Buffer + Luciferin Detection Reagent

*RLDL depozitare 24h la 22®C-25®C sau 3 luni la -30®C - -10®C (recomdare aliquotare)

Rezumat:

1) Tratament
2) H2O2 Substrate Solution, incubare 6h
3) ROS-Glo Solution, incubare 20 min
4) Citire luminescenta

***reactii abiotice care produc H2O2 (piruvati)

Experiment:

1) pregatire 80 ul mediu cu celule si tratament (ex: 70+10) (10.000 celule / well)

2) Diluare Substrat cu Dilution Buffer (10mM  125 ul ) *vortexare daca e neclar
3) adaugare 20 ul Substrat la mix(celule+tratament)=> 100ul/well, incubare 6h (Substrat 25uM)
* substrat nu mai mult de 6 ore in mediu
4) preparare Reconstituted Luciferin Detection Reagent = Reconstitution Buffer + Luciferin
Detection Reagent
* depozitare 24 ore la 22-25 grade sau 3 luni la -30- - grade
*recomandare aliquotare
5) inainte de utilizare se prepara ROS-Glo Detection Solution: 10 ul Cysteine si 10 ul Signal
Enhancer Solution la 1 ml Reconstituted Luciferin Detection Reagent
6) adaugare 100 ul/ well ROS-Glo, incubare 20 min
7) citire luminescenta