Plant Dehydrogenase (PDHA) Kit za ispitivanje aktivnosti
Bilješka: Uzmite dva ili tri različita uzorka za predviđanje prije testa.
Operativna oprema: Microplate Reader/ Spectrophotometer
Mačka ne: BC3125
Veličina:100T/48s
Komponente:
Reagens i: Powder×2. Skladištenje na 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.
Reagens II: Liquid 100 mL×2. Skladištenje na 4 ℃.
Reagens iii: Ethyl acetate, required but not provided.
Opis proizvoda:
The activity of plant dehydrogenase (PDHA) 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.
Potrebni reagensi i oprema, ali nisu pruženi:
Microplate Reader or spectrophotometer, vodena kupelji, stolna centrifuga, vodena kupelji, pipette, micro glass cuvette/96 well flat-bottom plate (non-polystyrene/polypropylene material), minobacač/homogenizator, ethyl acetate (express delivery is not allowed), led i destilirana voda.
Postupak:
Ja. Complex izvlačenje:
Collect 0.1 g tkiva, wash 3 ili 4 times with double steam water, blot dry with filter paper and set aside.
Ii. Odlučnost postupak:
- Preheat spectrophotometer/ microplate reader for 30 minuta, Podesite valnu duljinu na 485 NM, set zero with ethyl acetate.
- Add the following reagents in 5 mL EP tubes:
| Reagens | Contrast tube (Ac) | Epruveta (NA) |
| Uzorak (g) | 0.1 | 0.1 |
| Reagens i (ml) | – | 1 |
| Reagens 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. | ||
| Reagens iii (ml) | 1 | 1 |
Iii. Izračunavanje:
A. micro glass cuvette (light path of the cuvette, 1 cm)
Definicija jedinice: 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 bunar ploča (light path of the cuvette, 0.6 cm)
Definicija jedinice: 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: Vrijeme reakcije (h), 3 sati; W: Težina uzorka, g.
Bilješka
- 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.
- Uzmite dva ili tri različita uzorka za predviđanje prije testa. Dilute the supernatant if the absorbance is higher, Pomnožite razrijeđene vrijeme u formuli.
- If test with a 96 well flat-bottom plate, polystyrene, or polypropylene material 96 well flat-bottom plate is not recommended.
Eksperimentalni primjer:
- Weigh 1g aloe leaves, wash them with double distilled water for 3–4 times, absorb the water with filter paper, operate according to the determination steps, measure and calculate with 96 bunar ploča, ΔA=AT-AC= 0.244-0.146 = = 0.098, calculate the enzyme activity.
Dehydrogenase activity (U/g masa) = 66.7×ΔA÷W = 6.5366 U/g masa.
Povezani proizvodi：
BC2030/BC2035 Isocitrate Lyase (ICL) Kit za ispitivanje aktivnosti
BC3170/BC3175 Acetokinase (ACK) Kit za ispitivanje aktivnosti
BC2010/BC2015 Glycollic Oxidase Activity Assay Kit
Recenzije
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