Plant Dehydrogenase (Pdhi) Kit de test d'activité
Note: Prenez deux ou trois échantillons différents pour la prédiction avant le test.
Équipement d'exploitation: Microplate Reader/ Spectrophotometer
Chat non: BC3125
Taille:100T/48S
Composants:
Réactif I: Poudre × 2. Stockage à 4℃. Dissolve one bottle of powder in water to make 50 mL before use, prepare the solution when it will be use. Store at 4℃ and protect from light after prepared.
Réactif II: Liquide 100 mL×2. Stockage à 4℃.
Réactif III: Ethyl acetate, requis mais non fourni.
Description du produit:
The activity of plant dehydrogenase (Pdhi) is largely reflecting the active state of the organism, which can directly indicate the ability of biological cells to degrade its matrix.
The hydrogen acceptor 2,3,5-triphenyl tetrazolium chloride (TTC) generates red triphenylformazone (TFF) after receiving hydrogen during cell respiration. TFF has a characteristic absorption peak at 485 nm, the PDHA activity can quantified by measuring the absorbance at 485 nm.
Réactifs et équipements requis mais non fournis:
Microplate Reader or spectrophotometer, bain d'eau, centrifugeuse de bureau, bain d'eau, pipette, cuvette en verre micro/plaque à fond plat de 96 puits (non-polystyrene/polypropylene material), mortier/homogénéisateur, ethyl acetate (express delivery is not allowed), glace et eau distillée.
Procédure:
je. Complex extraction:
Collecter 0.1 g de tissu, laver 3 ou 4 times with double steam water, blot dry with filter paper and set aside.
II. Détermination procédure:
- Preheat spectrophotometer/ microplate reader for 30 minutes, ajuster la longueur d'onde à 485 nm, set zero with ethyl acetate.
- Ajouter les réactifs suivants dans 5 Tubes ML EP:
| Réactif | Tube de contraste (Ac) | Tube à essai (À) |
| Échantillon (g) | 0.1 | 0.1 |
| Réactif I (ml) | – | 1 |
| Réactif II (ml) | 2 | 1 |
| Mix thoroughly and stand in dark for 3 hours at 37℃, ice bath for 5 minutes immediately after take out. Discard the filtrate, blot dry the sample with filter paper, place in mortar / homogenizer. | ||
| Réactif III (ml) | 1 | 1 |
III. Calcul:
UN. micro glass cuvette (light path of the cuvette, 1 cm)
Définition de l'unité: One unit of enzyme activity is defined as the amount of enzyme increases the absorbance of every 0.01 for per hour every mg tissue protein in the reaction system.
PDHA Activity(U/g/h) =ΔA÷0.01÷T÷W=33×ΔA÷W
B. 96 bien plaque (light path of the cuvette, 0.6 cm)
Définition de l'unité: One unit of enzyme activity is defined as the amount of 1 mg of tissue increases the absorbance of every 0.005 for per hour in the reaction system.
PDHA Activity(U/g/h) =ΔA÷0.005÷T÷W=66.7×ΔA÷W
T: Temps de réaction (h), 3 heures; W: Poids de l'échantillon, g.
Note
- After prepared, Reagent I should store at 4℃, protect from light and used within one week. If it turns red, it cannot be used.
- Reagent III is volatile and toxic. For your health, please wear lab clothes, masks, and latex gloves.
- Ice bath to stop the reaction immediately after the dark reaction was completed. Discard the filtrate, blot dry the sample with filter paper as much as possible.
- Prenez deux ou trois échantillons différents pour la prédiction avant le test. Dilute the supernatant if the absorbance is higher, multiply dilute times in the formula.
- If test with a 96 assiette bien à fond plat, polystyrene, or polypropylene material 96 well flat-bottom plate is not recommended.
Exemple expérimental:
- Weigh 1g aloe leaves, wash them with double distilled water for 3–4 times, absorb the water with filter paper, fonctionner selon les étapes de détermination, measure and calculate with 96 bien plaque, ΔA=AT-AC= 0.244-0.146 = 0.098, calculate the enzyme activity.
Dehydrogenase activity (U/g masse) = 66.7×ΔA÷W = 6.5366 U/g masse.
Produits connexes：
BC2030/BC2035 Isocitrate Lyase (ICL) Kit de test d'activité
BC3170/BC3175 Acetokinase (Ack) Kit de test d'activité
BC2010/BC2015 Glycollic Oxidase Activity Assay Kit
Commentaires
Il n'y a pas encore de critiques.