Solarbio Malondialdehyde (MDA) Komplet za ispitivanje sadržaja

Malondialdehyde (MDA) Komplet za ispitivanje sadržaja

Bilješka: Potrebno je predvidjeti 2-3 Uzorci velike razlike prije formalnog određivanja. Operativna oprema: Spektrofotometar.

Mačka ne: BC0020

Veličina: 50T/48s

Komponente:

Komponenta	Type	Volume/Quantity	Skladištenje
Ekstrakcijski reagens	Liquid	60 ml × 1	4°C
Reagens i	Liquid	42 ml × 1	4°C
Reagens II	Powder	–	4°C
MDA working reagent	Liquid	20 mL Reagent I	4°C
		+ Reagens II
Reagens iii	Liquid	12 ml × 1	4°C

Bilješka: The working solution for MDA detection is difficult to dissolve, which can be heated at 70℃ and vibrated violently to promote dissolution. Or by ultrasonic treatment to promote dissolution.

Opis proizvoda:

• Lipid peroxide is generated through the interaction of oxygen free radicals with unsaturated fatty acids, eventually forming compounds such as malondialdehyde (MDA). The level of lipid peroxidation serves as an indicator, which can be assessed by measuring MDA levels.
• Under acidic and high-temperature conditions, a brown-red compound, 3,5,5-three methyl sulfamethoxazole-2,4-two ketone, is synthesized via a condensation reaction between MDA and thiobarbituric acid (TBA). This compound exhibits maximum absorption at 532 NM. The assessment of lipid peroxidation involves colorimetric analysis. Međutim, the presence of soluble sugars can interfere with the detection process. The reaction of soluble sugars with TBA produces a color reaction with absorption wavelengths at 450 nm and 532 NM. In this assay kit, the MDA content is determined by the variance in absorbance at 532 NM, 450 NM, i 600 NM.
• Due to the presence of sucrose in plant tissues and glucose in animal tissues, the kit includes two computational formulas tailored for sucrose and glucose. These formulas are suitable for lipid assessment.

Potrebni reagensi i oprema, ali nisu pruženi:

Spektrofotometar, vodena kupelji, stolna centrifuga, PRIJENOS,1 ML staklena kiveta, minobacač/homogenizator, led i destilirana voda.

Postupak:

Ja. Priprema uzorka:

Bakterije ili stanice:

Collect bacteria or cells into the centrifuge tube. 5 million bacteria or cells could be mixed with 1 ML reagensa za ekstrakciju. Use ultrasonication to split bacteria and cells (stavljen na led, ultrasonic power 200W, ultrasonic time 3 sekundi, interval 10 sekundi, ponoviti za 30 vrijeme). Centrifuga na 8000 × g za 10 minuta u 4 ℃ za uklanjanje netopljivih materijala, and take supernatant on ice before testing.

Tkivo:

0.1 g of tissue could be mixed with 1 mL of Extraction reagent and fully homogenized on ice bath. Then centrifuge at 8000 × g za 10 minuta u 4 ℃ za uklanjanje netopljivih materijala, i prije testiranja uzmite supernatant na ledu.

Serum:

Detect

Ii. Odlučnost postupak:

1. Preheat spectrophotometer for more than 30 minuta, set zero with distilled
2. Dodajte reagense sa sljedećim popisom:

Reagens (μL)	Epruveta (T)	Prazna cijev (B)
MDA working reagent	600	600
Uzorak	200	–
Destilirana voda	–	200
Reagens ⅲ	200	200

The mixture would be incubated at 100℃ for 60 minuta (tightly close to prevent moisture loss), cooled on ice, and centrifuged at 10000 × g za 10 minutes at room temperature to remove insoluble materials. Take supernatant in 1 ML staklena kiveta, and measure the absorbance at 450 NM, 532 nm and 600 NM.

∆A450=A450(T)-A450(B), ∆A532=A532(T)-A532(B), ∆A600 =A600(T)-A600(B). Blank tube needs to test once or twice.

Iii. Izračunavanje:

1. Tkivo, Bakterije ili kultivirane stanice

• Koncentracija proteina:

MDA (nmol/mg prot)= =(6.45×(∆A532-∆A600)-1.29×∆A450)×Vrv÷(Cpr×Vs)

=5×(6.45×(∆A532-∆A600)-1.29×∆A450)÷Cpr

• Težina uzorka:

MDA (nmol/g weight)= =( 6.45×(∆A532-∆A600)- 1.29×∆A450)×Vrv÷(W × vs ÷ vsv)

=5×(6.45×(∆A532-∆A600)- 1.29×∆A450)÷W

• Cellamount:

MDA (nmol/104cell)= =( 6.45 ×(∆A532-∆A600)- 1.29×A∆450)×Vrv÷(400× vs ÷ vsv)

=0.01×(6.45×(∆A532-∆A600)- 1.29×∆A450)

• Serum:

MDA (nmol/mL)= =(6.45×(ΔA532 -ΔA600)-1.29×ΔA450)×Vrv÷Vs

=5×(6.45×(ΔA532 -ΔA600)-1.29×ΔA450)

2. Plants tissue:

• Težina uzorka

MDA (nmol/g weight)= = (6.45×(∆A532-∆A600)-0.56×∆A450)×Vrv÷(W × vs ÷ vsv)

=5×(6.45 ×(∆A532-∆A600)- 0.56×∆A450)÷W

• Koncentracija proteina:

MDA (nmol/mg prot)= =( 6.45 ×(∆A532-∆A600)- 0.56×∆A450)×Vrv÷(Cpr×Vs)

=5×(6.45 ×(∆A532-∆A600)- 0.56×∆A450)÷Cpr

Vrpca: Ukupni volumen reakcije, 1 ml; Vs: Volumen uzorka, 0.2 ml;

VSV: Valum ekstrakcije, 1 ml;

CPR: Koncentracija proteina uzorka, mg/ml; W: Težina uzorka, g;

400: Ukupni broj bakterija i stanica, 5 milijun.

Bilješka:

If it is found that the absorbance value of the sample is too low, the boiling water bath time can be adjusted from 60 minutes to 90 minutes or longer. The detection of MDA in the same experiment needs to be extended to the same time to avoid errors.

Reference

• Spitz D R, Oberley L W. An assay for superoxide dismutase activity in mammaliantissue homogenates[J]. Analitička biokemija,1989
• Masayasu M, Hiroshi y. Pojednostavljena metoda ispitivanja superoksidne aktivnosti dismutaze za kliničku upotrebu[J]. Clinica Chimica

Povezani proizvodi:

BC3590/BC3595 Hydrogen Peroxide (H2O2) Komplet za ispitivanje sadržaja

BC1090/BC1095 Xanthine Oxidase (XOD) Kit za ispitivanje aktivnosti

BC0690/BC0695 Glucose Oxidase (GOD) Activity Assay Kit BC1270/BC1275 Protein Carbonyl Content Assay Kit BC1280/BC1285 Diamine Oxidase (DAO) Activity Assay Kit BC1290/BC1295 Superoxide Anion Content Assay Kit

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