Thiobarbituric acid reactants (TBARS) Colorimetric Assay Kit

    • Biochemical Assay Kit-Elabscience
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    • Biochemical Assay Kit-Elabscience
    • Biochemical Assay Kit-Elabscience

      Catalog number:E-BC-K298-M

      Size:
      • 48T
      • 96T
      Qty:
      - +
      Price: $180

      Detection method: Colorimetric method

      Detection instrument: Microplate reader

      Valid period: 6 months

      Lead Time: 7~10 daysWelcome to order from local distributors.

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      General information

      Detection significance

      The organism produces oxygen free radicals through enzyme system and non-enzyme system, attacks polyunsaturated fatty acids in biofilm, induces lipid peroxidation, and thus forms lipid peroxide. Malondialdehyde (MDA) is one of the common products of lipid peroxidation in organisms. In clinical science, MDA is a biomarker of lipid peroxidation, which can reflect the degree of lipid peroxidation in organism and indirectly reflect the degree of cell injury. 

      Detection principle

      TBARS and TBA can react under high temperature and acid conditions and then form a pink compound, the concentration of which is linearly related to the concentration of TBARS in the sample. The TBARS concentration can be calculated by measuring the OD values at 530-540 nm.

      The key point

      1.  The temperature should be controlled at 95~100℃ when incubate in water bath for 1 hour.

      2.  In water bath reaction, do not cover the caps of tubes tightly. It is recommended to seal the test tube mouth with the preservative film and make a hole in the preservative film.

      3.  Prevent the formulation of bubbles when the supernatant is transferred into the microplate.

      Operation procedures

      Plate set up

       

      1

      2

      3

      4

      5

      6

      7

      8

      9

      10

      11

      12

      A

      A

      A

      S1

      S9

      S17

      S25

      S33

      S41

      S49

      S57

      S65

      S73

      B

      B

      B

      S2

      S10

      S18

      S26

      S34

      S42

      S50'

      S58

      S66

      S74

      C

      C

      C

      S3

      S11

      S19

      S27

      S35

      S43

      S51

      S59

      S67

      S75

      D

      D

      D

      S4

      S12

      S20

      S28

      S36

      S44

      S52

      S60

      S68

      S76

      E

      E

      E

      S5

      S13

      S21

      S29

      S37

      S45

      S53

      S61

      S69

      S77

      F

      F

      F

      S6

      S14

      S22

      S30

      S38

      S46

      S54

      S62

      S70

      S78

      G

      G

      G

      S7

      S15

      S23

      S31

      S39

      S47

      S55

      S63

      S71

      S79

      H

      H

      H

      S8

      S16

      S24

      S32

      S40

      S48

      S56

      S64

      S72

      S80

                                  [Note]:A-H:standard wells;S1-S80:sample wells.

      The dilution of standard curve

      Dilute 200 μmol/L Standard Solution with double distilled water to a serial concentration. The recommended dilution gradient is as follows: 100, 80, 60, 40, 20, 10, 5, 0 μmol/L.

      Operation steps

      1)      Standard tube: Take 0.1 mL of Standard solution with different concentrations into numbered 10 mL glass tubes.

         Sample tube: Take 0.1 mL of tested Sample into numbered 10 mL glass tubes.

      2)      Add 0.1 mL of Reagent 1 into each tube of Step 1.

      3)      Add 4 mL of Chromogenic agent into each tube of Step 2.

      4)      Fasten the tube mouth with fresh-keeping film, mix fully, and make a small hole in the film. Then incubate the tubes at 100℃ for 1 hour.

      5)      Take the tubes out and put them in an ice bath to stop the reaction. After incubation on ice for 10 min, centrifuge the tubes at 1600 × g for 10 min.

      6)      Take 0.25 mL the supernatant to the Microplate with a micropipette (the precipitation cannot be added to the Microplate).

      7)      Measure the OD value at 532 nm with Microplate Reader. 

      Operation table

       

      Standard tube

      Sample tube

      Standard solution of different concentrations (mL)

      0.1

       

      Sample (mL)

       

      0.1

      Reagent 1 (mL)

      0.1

      0.1

      Chromogenic agent (mL)

      4

      4

      Fasten the tube mouth with fresh-keeping film, mix fully, and make a small hole in the film. Then incubate the tubes at 100 for 1 hour. After incubation on ice for 10 min, centrifuge the tubes at 1600 × g for 10 min. Take 0.25 mL the supernatant to the Microplate with a micropipette. Determine the OD at 532 nm with Microplate Reader.

      Performance characteristics

      Technical parameter

      Detection range 2.6-100 μmol/L Average inter-assay CV 7.1%
      Sensitivity 0.85 μmol/L Average intra-assay CV 4.3%
      Average recovery rate 100.9%

      Standard curve

      Dilute 200 μmol/L Standard Solution with double distilled water to a serial concentration. The recommended dilution gradient is as follows: 100, 80, 60, 40, 20, 10, 5, 0 μmol/L. Then carry the assay according to the operation table.


      Plot the standard curve by using OD value of standard and correspondent concentration as y-axis and x-axis respectively. Create the standard curve with graph software (or EXCEL). The concentration of the sample can be calculated according to the formula based on the OD value of sample. The standard curve is: y= ax + b.


      Example analysis

      For mouse liver tissue, take 0.1 mL of sample, carry the assay according to the operation table.

      The results are as follows:

      standard curve: y = 0.0038 x – 0.0013, the average OD value of the sample well is 0.106, the average OD value of the blank well is 0.047, the concentration of protein in sample is 15.18 gprot/L, and the calculation result is:

      TBARS content (μmol/gprot)=(0.106-0.047+0.0013)÷0.0038÷15.18=1.05μmol/gprot

      Detect rat serum, rat plasma, 10% mouse liver tissue homogenate (the concentration of protein is 15.18 gprot/L), 10% mouse heart tissue homogenate (the concentration of protein is 5.55 gprot/L) according to the protocol, the result is as follows:

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