Solarbio Malondialdéhyde (MDA) Kit d'analyse de contenu

Malondialdéhyde (MDA) Kit d'analyse de contenu

Note: Il faut prévoir 2-3 échantillons de grande différence avant la détermination formelle. Équipement d'exploitation: Spectrophotomètre.

Chat non: BC0020

Taille: 50T/48S

Composants:

Note: 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.

Description du produit:

• 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. Cependant, 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, et 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.

Réactifs et équipements requis mais non fournis:

Spectrophotomètre, bain d'eau, desk centrifuge, pipette de transfert,1 Cuvette en verre ml, mortier/homogénéisateur, glace et eau distillée.

Procédure:

je. La préparation des échantillons:

Bactéries ou cellules:

Collect bacteria or cells into the centrifuge tube. 5 million bacteria or cells could be mixed with 1 mL de réactif d'extraction. Use ultrasonication to split bacteria and cells (placé sur la glace, ultrasonic power 200W, ultrasonic time 3 secondes, interval 10 secondes, répéter pour 30 fois). Centrifuger à 8000 ×g pour 10 minutes à 4℃ pour éliminer les matières insolubles, and take supernatant on ice before testing.

Tissu:

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 pour 10 minutes à 4℃ pour éliminer les matières insolubles, et prendre le surnageant sur la glace avant de tester.

Sérum:

Détecter

II. Détermination procédure:

1. Preheat spectrophotometer for more than 30 minutes, mettre à zéro avec distillé
2. Ajoutez des réactifs avec la liste suivante:

The mixture would be incubated at 100℃ for 60 minutes (tightly close to prevent moisture loss), cooled on ice, and centrifuged at 10000 ×g pour 10 minutes at room temperature to remove insoluble materials. Take supernatant in 1 Cuvette en verre ml, 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. Calcul:

1. Tissu, bactéries ou cellules cultivées

• Concentration en protéines:

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

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

• Poids de l'échantillon:

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

=5×(6.45×(∆A532-∆A600)- 1.29×∆A450)÷W

• Montant de la cellule:

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

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

• Sérum:

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

=5×(6.45×(ΔA532 -ΔA600)-1.29×ΔA450)

2. Plants tissue:

• Poids de l'échantillon

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

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

• Concentration en protéines:

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

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

Corde: Volume total de réaction, 1 ml; Contre: Volume d'échantillon, 0.2 ml;

Vsv: Volume d'extraction, 1 ml;

Cpr: Concentration de protéines de l'échantillon, mg/ml; W: Poids de l'échantillon, g;

400: Nombre total de bactéries et de cellules, 5 million.

Note:

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 pour 90 minutes or longer. The detection of MDA in the same experiment needs to be extended to the same time to avoid errors.

Les références

• Spitz D R, Oberley L W. An assay for superoxide dismutase activity in mammaliantissue homogenates[J.]. Biochimie analytique,1989
• Masayasu M., Hiroshi Y.. Une méthode de dosage simplifiée de l'activité de la superoxyde dismutase à usage clinique[J.]. Clinica Chimica

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