FLUOR DE LYS®SIRT3 fluorometric drug discovery assay kit

Store all components except the microplates and instruction booklet at -70°C for the highest stability. The SIRT3 enzyme, Prod. No. BML-SE270, must be handled with particular care in order to retain maximum enzymatic activity. Defrost it quickly in a RT water bath or by rubbing between fingers, then immediately store on an ice bath. The remaining unused extract should be refrozen quickly, by placing at -70°C. If possible, snap freeze in liquid nitrogen or a dry ice/ethanol bath. To minimize the number of freeze/thaw cycles, aliquot into separate tubes and store at -70°C. The 5x Developer II (^Prod. No. BML-KI176) can be prone to precipitation if thawed too slowly. It is best to thaw this reagent in a room temperature water bath and, once thawed, transfer immediately onto ice.

SIRT3 (Sirtuin 3) (human, recombinant) (Prod. No. BML-SE270) (500 U, one U=1 pmol/min at 37°C, 500μM, FLUOR DE LYS ® -SIRT2 substrate (Prod. No. BML-KI179), 500μM NAD ; Recombinant enzyme dissolved in 25mM TRIS, pH 7.5, 100mM sodium chloride, 5mM dithiothreitol and 10% glycerol. See vial label for activity and protein concentrations) Storage: -70°C, avoid freeze/thaw cycles! FLUOR DE LYS ® -SIRT2, Deacetylase substrate (Prod. No. BML-KI179) (100μl; 5mM solution in 25mM TRIS/Cl, pH 8.0, 137mM sodium chloride, 2.7mM potassium chloride, 1mM magnesium chloride) . Note: this SIRT2 substrate is also hydolyzed by SIRT3. FLUOR DE LYS ® Developer II Concentrate (5x) (Prod. No. BML-KI176) (5 x 250 μL; 5x Stock Solution; Dilute in Assay Buffer before use) Storage: -70°C NAD (Sirtuin Substrate) (Prod. No. BML-KI282) (500 μl; 50 mM β-Nicotinamide adenine dinucleotide (oxidized form) in 50mM TRIS/Cl, pH 8.0, 137mM sodium chloride, 2.7mM potassium chloride, 1mM magnesium chloride) Storage: -70°C Nicotinamide (Sirtuin Inhibitor) (Prod. No. BML-KI283) (500μl; 5 mM Nicotinamide in 50mM TRIS/Cl, pH 8.0, 137mM sodium chloride, 2.7mM potassium chloride, 1mM magnesium chloride) Storage: -70°C Suramin sodium (Sirtuin Inhibitor) (Prod. No. BML-KI285) (10 mg; Solid MW: 1429.2, soluble in water or assay buffer (to 25mM) ) Storage: -70°C FLUOR DE LYS ® Deacetylated Standard (Prod. No. BML-KI142) (30 μl; 10mM in DMSO) Storage: -70°C Sirtuin Assay Buffer (Prod. No. BML-KI286) (20ml; 50mM TRIS/Cl, pH 8.0, 137mM sodium chloride, 2.7mM potassium chloride, 1mM magnesium chloride, 1 mg/ml bovine serum albumin) Storage: -70°C 1/2 volume microplate (Prod. No. BML-KI101) Storage: Room temperature 1/2 volume white microplate (Prod. No. BML-KI110) Storage: Room temperature