CoenzymeⅠNAD(H) Content Assay Kit (WST colorimetry)

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辅酶Ⅰ NAD(H)含量检测试剂盒（WST显色法）

Cat:

NA0173

Assay Type:

Spectrophotometer

Brand:

sunlong medical

Specificity:

50T/24S

Storage instructions：

-20℃

Product Overview：

Components:

Acid Extract solutiont : Liquid 15 mL×1. Storage at 2-8℃.

Alkaline Extract solutiont: Liquid 15 mL×1. Storage at 2-8℃.

Reagent I : Liquid 20 mL×1. Storage at 2-8℃.

Reagent II : Liquid 6 mL×1. Storage at 2-8℃.

Reagent III : Liquid 12 mL×1. Storage at 2-8℃..

Reagent IV: Liquid 3 mL×1. Storage at -20℃.

Reagent V : Liquid 40 mL×1. Storage at 2-8℃.

NAD standard: Powder×1. Storage at -20℃.Add 1.5 mL of distilled water before use to obtain a

standard of 2 µmol/mL, which can be stored at -20°C for 2 weeks.

NADH standard: Powder×1. Storage at -20℃.Add 1.4 mL of distilled water before use to obtain a

standard of 2 µmol/mL, which can be stored at -20°C for 2 weeks.

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Unit:

Price: $121

Product PDFs

Datasheet:

Other Information
Citations
Protein Attributes
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Coenzyme I, including reduced and oxidized forms, plays a role in transferring hydrogen in biological

oxidation. Oxidized coenzyme I, also known as nicotinamide adenine dinucleotide (NAD⁺), is a coenzyme

of dehydrogenase, which plays an irreplaceable role in glycolysis, gluconeogenesis, tricarboxylic acid

cycle and respiratory chain. The intermediate product will pass off the hydrogen to NAD, making it NADH

(reduced coenzyme I). NADH acts as a hydrogen carrier to synthesize ATP by chemical osmotic coupling

in the respiratory chain. NAD(H) has important physiological significance in the body, and is closely

related to material metabolism, energy metabolism, anti-aging, anti-oxidation and the occurrence of some

diseases. Decreased levels of coenzyme I in the body can lead to cell damage or decline.

The NAD⁺and NADH in the samples were extracted with acidic and alkaline extracting solutions,

respectively. Under the action of 1-mPMS, WST-1 can react with NADH to produce water-soluble

formazan, which has a characteristic absorption peak at 450 nm, while NAD⁺can be dehydrogenated by

ethanol The enzyme was reduced to NADH, which was further detected by WST-1.

Reagents and Equipment Required but Not Provided:

Spectrophotometer, centrifuge, water bath, mortar/homogenizer, sonicator, adjustable pipette, 1 mL

glass cuvette, ice, and distilled water.

Sample Preparation：

1. Serum

Extract NAD⁺: Take 0.1 mL of serum (slurry), add 0.5 mL of acidic extract, boil for 5 minutes (cover

tightly to prevent water loss), and after cooling in an ice bath, centrifuge at 10,000g at 4°C for 10

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minutes; take 200 µL of supernatant, add an equal volume of alkaline extract ; Mix well, centrifuge at

10,000g at 4°C for 10 min, take the supernatant, and store on ice for testing.

Extract NADH: Take 0.1 mL of serum (slurry), add 0.5 mL of alkaline extract, boil for 5 minutes

(cover tightly to prevent water loss), after cooling in an ice bath, centrifuge at 10,000g at 4°C for 10

minutes, take 200 µL of supernatant, and add an equal volume of acidic extract ; Mix well, centrifuge

at 10000g at 4°C for 10min, take the supernatant and store on ice for testing.

2. Tissue

Extract NAD⁺: Weigh about 0.1g of tissue, add 0.5mL of acidic extract, grind in ice bath, boil for

5min (cover tightly to prevent water loss), after cooling in ice bath, centrifuge at 10,000g at 4°C for

10min, take 200µL of supernatant, add an equal volume of alkaline extract ; Mix well, centrifuged at

10,000 g at 4°C for 10 min, and the supernatant was taken and stored on ice for testing.

Extract NADH: Weigh about 0.1 g of tissue, add 0.5 mL of alkaline extraction solution, grind in an

ice bath, boil for 5 min (cover tightly to prevent water loss), after cooling in an ice bath, centrifuge at

10,000 g at 4°C for 10 min, take 200 µL of supernatant, and add an equal volume of acidic extract ;

Mix well, centrifuged at 10,000 g at 4°C for 10 min, and the supernatant was taken and stored on ice

for testing.

3. Bacteria or cells

Extract NAD⁺: Collect 5 million cells or bacteria, add 0.5mL of acidic extract, ultrasonically disrupt

for 1min (intensity 20% or 200W, ultrasonic for 2s, stop for 1s), boil for 5min (cover tightly to prevent

water loss), cool in ice bath, 10000g centrifuge at 4°C for 10min, take 200uL of the supernatant into

another new centrifuge tube, add an equal volume of alkaline extract to neutralize, mix well,

centrifuge at 10,000g at 4°C for 10min, take the supernatant and store it on ice for testing .

Extract NADH: Collect 5 million cells or bacteria, add 0.5mL alkaline extract, ultrasonically disrupt

for 1min (intensity 20% or 200W, ultrasonic for 2s, stop for 1s), boil for 5min (cover tightly to prevent

water loss), cool in an ice bath, Centrifuge at 10,000g at 4°C for 10min, take 200uL of the supernatant

into another new centrifuge tube, add an equal volume of acidic extract to neutralize, mix well,

centrifuge at 10,000g at 4°C for 10min, take the supernatant, and store it on ice for testing .

Determination

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