Diamine oxidase test kit

Diamine oxidase test kitMicro method

Attention: Before formal testing, select 2-3 samples with significant expected differences for prediction.

Item No.BC1285

Specifications100T/48S

Product Content:

Extraction solution: liquid100mL×1Bottle,4℃保存。

Reagent 1: Liquid0.25mL×1Support,4℃保存。

Reagent 2: Liquid2mL×1Pipe,4℃保存。

Reagent 3: Liquid1mL×1Pipe,4℃保存。

Product Description:

DAO(EC1.4.3.6)Widely present in animals (intestinal mucosa, lungs, liver, kidneys, etc.), plants, and microorganisms. Catalytic oxidation of polyamines to aldehydes, whose activity is closely related to nucleic acid and protein synthesis, can reflect the integrity and degree of damage to the intestinal mechanical barrier.

DAOCatalyze cadaverine to produce aldehydes and *, and add excessive horseradish peroxidase externally to catalyze * oxidation of neighboring compounds

Phenylamine generates oxidized ortho phenylamine in460nmThere is a characteristic absorption peak at this wavelength. By measuring the rate of increase in absorbance at this wavelength, calculateDAOActivity.

Self provided experimental supplies and instruments:

Balance, low-temperature centrifuge, visible spectrophotometer/ELISA reader, trace quartz colorimetric dish/96Orifice plate, distilled water.

Operation steps:

1、 Extraction of crude enzyme solution:

1. Organization: According to organizational quality（g）Volume of extraction solution(mL)for1：5~10The proportion of (recommended to be around)0.1gOrganize, join1mLExtract the liquid and homogenize it in an ice bath, then10000g，4℃ centrifugation20minTake the supernatant and place it on ice for testing.

2. Bacteria and fungi: according to the number of cells（10⁴Individual: volume of extraction solution（mL）For500~1000：1The proportion (suggested)500Ten thousand cells added1mLExtraction solution), ice bath ultrasonic disruption of cells (power)300w, Ultrasound3Seconds, intervals7Second, total time3min）； then10000g，4℃, centrifugation10minTake the supernatant and place it on ice for testing.

3. Serum and other liquids: direct measurement.

2、 Measurement operation table:

1Spectrophotometer/Preheating of ELISA reader30minAdjust the wavelength460nmAdjust distilled water to zero.

2Operation table

3、 Enzyme activity calculation formula:

a.The calculation formula for measuring with a trace quartz colorimetric dish is as follows

（1）Calculated based on sample protein concentration:

Definition of enzyme activity: inpH7.2The temperature is37Under ℃ conditions, every milligram of tissue protein catalyzes production per minute1nmol H₂O₂The required enzyme quantity is one enzyme activity unit（U）.

DAOActivity（nmol/min/mg prot）=A₄₆₀÷（ε×d）×VAnti General÷（Vsample×Cpr）÷T= 18×A 460 ÷Cpr

（2）Calculated by sample quality:

Definition of enzyme activity: inpH7.2The temperature is37Under ℃ conditions, catalytic production occurs per gram of tissue per minute1nmol H₂O₂The required enzyme quantity is defined as one unit of enzyme activity.

DAOActivity（nmol/min/g）=A₄₆₀÷（ε×d）×VAnti General÷（Vsample÷VSample General×W）÷T= 18×A 460 ÷W

（3）Calculated by the number of cells:

Definition of enzyme activity: inpH7.2The temperature is37Under the condition of ℃, every10⁴Cells catalyze production every minute1nmol H²O²The required enzyme quantity is defined as one unit of enzyme activity.

DAOActivity（nmol/min/10⁴cell）=A₄₆₀÷（ε×d）×VAnti General÷（Vsample÷VSample General×Number of cells÷T= 18×A₄₆₀÷cell count

(4)Calculated by liquid volume

Definition of enzyme activity: inpH7.2The temperature is37Under the condition of ℃, every milliliter of serum catalyzes production per minute1nmol H₂O₂The required enzyme quantity is defined as one unit of enzyme activity.

DAOActivity（nmol/min/mL）=A₄₆₀÷（ε×d）×VAnti General÷Vsample÷T= 18×A₄₆₀

εOxidation type ortho fennel amine millimolar extinction coefficient:7.5 L/mmol/cm；dColor dish optical diameter,1cm；VAnti General: AntiThe total volume should be,0.2mL；VSample: Sample volume in the reaction,0.05mL；VSample total: Add the volume of extraction solution,1mL；Cpr：

Sample protein concentration,mg/mL；W: Sample quality,g；TResponse time,30min

b.use 96 The calculation formula for orifice plate measurement is as follows

（1）Calculated based on sample protein concentration:

Definition of enzyme activity: inpH7.2The temperature is37Under ℃ conditions, every milligram of tissue protein catalyzes production per minute1nmol H₂O₂The required enzyme quantity is one enzyme activity unit（U）.

DAOActivity（nmol/min/mg prot）=A₄₆₀÷（ε×d）×VAnti General÷（Vsample×Cpr）÷T= 36×A 460 ÷Cpr

（2）Calculated by sample quality:

Definition of enzyme activity: inpH7.2The temperature is37Under ℃ conditions, catalytic production occurs per gram of tissue per minute1nmol H₂O₂The required enzyme quantity is defined as one unit of enzyme activity.

DAOActivity（nmol/min/g）=A₄₆₀÷（ε×d）×VAnti General÷（Vsample÷VSample General×W）÷T= 36×A₄₆₀÷W

（3）Calculated by the number of cells:

Definition of enzyme activity: inpH7.2The temperature is37Under the condition of ℃, every10⁴Cells catalyze production every minute1nmol H₂O₂The required enzyme quantity is defined as one unit of enzyme activity.

DAOActivity（nmol/min/10⁴cell）=A₄₆₀÷（ε×d）×VAnti General÷（Vsample÷VSample General×Number of cells÷T = 36×A₄₆₀÷cell count

(4)Calculated by liquid volume

Definition of enzyme activity: inpH7.2The temperature is37Under the condition of ℃, every milliliter of serum catalyzes production per minute1nmol H₂O₂The required enzyme quantity is defined as one unit of enzyme activity.

DAOActivity（nmol/min/mL）=A₄₆₀÷（ε×d）×VAnti General÷Vsample÷T= 36×A₄₆₀

εOxidation type ortho fennel amine millimolar extinction coefficient:7.5 L /mmol/cm；d：96 Aperture plate optical diameter,0.5cm；VAnti General:

Total reaction volume,0.2mL；VSample: Sample volume in the reaction,0.05mL；VSample total: Add the volume of extraction solution,1mL；Cpr: Sample protein concentration,mg/mL；W: Sample quality,g；TResponse time,30min

Notes:

1IfODValue less than0.01Appropriately increase the sample size for extraction;ODValue greater than0.8The crude enzyme solution can be diluted appropriately or the sample size for extraction can be reduced.

2The protein content of the sample needs to be measured separately.

Diamine oxidase test kitOrdering instructions:

1. The vast majority of products are in stock and can generally be ordered and shipped on the same day.
2. Some non essential products require a 1-2 day advance booking, while overseas futures require a 3-6 week advance booking.
3. Some product prices may change due to factors such as delivery time and batch. If there are any changes, the new price on the day of ordering shall prevail.
4. The daily order deadline is 16:00 sharp, and in some cities it can be until 17:00 sharp. If the deadline is exceeded and the shipment cannot be made on the same day, it will be arranged for the next day.
5. After processing the payment, please notify us by fax, text message, etc. of the receipt and the recipient's address, name, etc.

Transportation instructions:

Ultra low temperature products: Dry ice is added during the transportation of extremely low temperature products. Wrap the product with dry ice, seal it with a foam box, glue the foam box layer by layer with adhesive tape, put it into the box of SOLEBAR, and then deliver it to the cooperative express delivery, which will deliver it safely, quickly, efficiently, and confidently to the customers, ensuring the stability of the product's nature and escorting your experiment.
Low temperature products: During the transportation of low-temperature products, add Solaibao ice packs for transportation. Wrap the product with ice bags in advance, then seal it with foam boxes, seal the foam boxes with adhesive tape, and put it into the boxes of SOLEBAR (the heat preservation effect can last for a week if it is less), and then deliver it to the cooperative express, which can deliver it safely, quickly, efficiently and confidently to the customers, ensuring the stability of the product properties and escorting your experiment.
Normal temperature products: Normal temperature products do not require ice or special packaging during transportation. The products are quickly distributed by the company's warehouse personnel, and accurate, fast, and efficient express delivery ensures that the products are delivered to you quickly.