Materials and Reagents

1. Sea sand (e.g. Merck, catalog number: 107711 ) to facilitate mechanically homogenizing the samples
   
2. Liquid nitrogen
   
3. Horse radish peroxidase (HRP) (e.g. Sigma-Aldrich, catalog number: P8375 )
   
4. Urate oxidase (UOX) (e.g. Sigma-Aldrich, catalog number: U0880 )
   
5. Xanthine oxidase (XO) (e.g. SERVA Electrophoresis GmbH, catalog number: 38418 )
   
6. Amplex^® ultra red (AR) (Life Technologies, Invitrogen^TM, catalog number: A36006 )
   
7. Uric acid (e.g. Sigma-Aldrich, catalog number: U2625 )
   
8. Xanthine (e.g. Sigma-Aldrich, catalog number: X7375 )
   
9. Dimethylsulfoxide (DMSO) (e.g. Sigma-Aldrich, catalog number: D4540 )
   
10. Prepare 10 mM AR-stocks (see Recipes)
11. Prepare stocks of HRP, UOX and XO (see Recipes)
    

Equipment

1. Standard flat-bottom microplates (e.g. Greiner, catalog number: 655161 )
   
2. Spectrophotometer equipped for reading multiwell plates (e.g. MultiSkan Go, Thermo Fisher Scientific)
   
3. Heating block with shaker function
   
4. Ultracentrifuges for 20,000 x g (and ideally for 40,000 x g)
   
5. Rotator for grinding samples (e.g. Heidolph RZR 2020, but a simpler device may also do) with a tissue-grinder which fits neatly into a conically-tapered 1.5 ml tube (see Figure 1).
   
   
   Figure 1. Tissue grinder for microtubes (Taylor Scientific)
   
   
6. Precision analytic balance, e.g. Mettler Toledo XS series, ideally equipped with an ErgoClip holder for 1.5 ml tubes
   
7. Heated magnetic stirrer
   
8. Micropipettes (1,000, 200, 100, 20 µl)
   
9. Multi-channel micropipette (range 50-200 µl)
   
10. 1.5 ml reaction tubes
    
11. 0.5 ml reaction tubes
    
12. 15 ml tubes (if dilutions of extracts become necessary)
    

Procedure