Protein Carbonyl Colorimetric Assay Kit (Tissue and serum samples)

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

      Catalog number:E-BC-K117

      Size:
      • 50 Assays
      • 100 Assays
      Qty:
      - +
      Price: $240

      Detection method: Colorimetric method

      Detection instrument: Spectrophotometry (UV)

      Valid period: 6 months

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

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      Application

      This kit can be used for detection of protein carbonyl content in serum (plasma), tissue, hydrothorax, cell supernatant, and other samples. This kit (100 Assays) can detect 50 samples.

       

      Detection significance

      The reactive oxygen species produced by the aerobic metabolism in the body can cause the oxidation of DNA, lipid and protein. The secondary reaction of the amino acid side chain of the protein with the lipid oxidation product is the main cause of the formation of the carbonyl. Carbonyl is a biological marker of ROS-mediated protein oxidation.

       

      Detection principle

      The content of protein carbonyl increased after oxidation, and the carbonyl group reacted with 2, 4-dinitrophenylhydrazine to form a reddish brown precipitate. The absorbance can be measured at 370 nm after the precipitation is dissolved. The carbonyl content can be calculated indirectly.

                                           



      Experimental instruments

      Test tube, Micropipettor, Vortex mixer, Centrifuge, 37 water bath, Spectrophotometer (370 nm)


      Preparation of sample

      1. Serum (Plasma), hydrothorax, cell supernatant: Detect the sample directly.

      2. Tissue sample: Accurately weigh the tissue sample, add the Reagent 1 at the ratio of Weight (g): Volume (mL) = 1:9. Mechanically homogenize the sample in ice water bath to prepare 10% homogenate. Centrifuge at 10000 g for 10 min, then take 450 μL the supernatant and add 50 μL of Reagent 2 application solution. Stand for 10min at room temperature, centrifuge at 11580 g for 10 min and take the supernatant for test.

       

      Operation steps

       

      Sample tube

      Control tube

      Sample (mL)

      0.1

      0.1

      Reagent 3 (mL)

      0.4

       

      Reagent 4 (mL)

       

      0.4

      Mix fully by swirling for 1 min, react with shading light at 37 for 30 min accurately.

      Reagent 5 (mL)

      0.5

      0.5

      Mix fully by swirling for 1 min, centrifuge at 13780 g for 10 min at 4, discard the supernatant and keep the precipitate.

      Anhydrous ethanol-ethyl acetate mixture application solution (mL)

      1.0

      1.0

      Mix fully by swirling for 1 min, centrifuge at 13780 g for 10 min at 4, discard the supernatant and keep the precipitate.

      Anhydrous ethanol-ethyl acetate mixture application solution (mL)

      1.0

      1.0

      Mix fully by swirling for 1 min, centrifuge at 13780 g for 10 min at 4, discard the supernatant and keep the precipitate.

      Anhydrous ethanol-ethyl acetate mixture application solution (mL)

      1.0

      1.0

      Mix fully by swirling for 1 min, centrifuge at 13780 g for 10 min at 4, discard the supernatant and keep the precipitate.

      Anhydrous ethanol-ethyl acetate mixture application solution (mL)

      1.0

      1.0

      Mix fully by swirling for 1 min, centrifuge at 13780 g for 10 min at 4, discard the supernatant and keep the precipitate.

      Reagent 6 (mL)

      1.25

      1.25

      Mix fuuly and incubate at 37 water bath for 15 min accurately.

      Mix fully by swirling to dissolve the precipitate fully. Centrifuge at 12000rpm for 15 min, then take the supernatant. Set to zero with Reagent 6 and measure the OD values of each tube at 370 nm with 0.5 cm diameter quartz cuvette. Meanwhile, determine the protein concentration of supernatant (E-BC-K318, E-BC-K168, E-BC-K165).


      Technical parameter

      1. The sensitivity of the kit is 0.02 nmol/L.

      2. The intra-CV is 4.5% and the inter CV is 8.6%.

      3. The recovery of the kit is 101%.

      4. The detection range of the kit is 0.02-10 nmol/L.

       

      Note

      1. The kit is for scientific research only.

      2.  Instructions should be followed strictly, changes of operation may result in unreliable results.

      3. The validity of kit is 6 months.

      4. Do not use components from different batches of kit.

      5. When washing the precipitate with anhydrous ethanol-ethyl acetate mixture application solution, the vortex must be voilent the mixing time should not be less than 1 min and the precipitate must be washed to white. If the precipitate still appear yellow, increase the washing times properly of anhydrous ethanol-ethyl acetate mixture application solution to ensure the washing process is sufficient. Otherwise the result will be higher.

      6. The speed of centrifuge should not be reduced, otherwise the result will be higher.

      7. It is recommended that the round bottom test tube instead of the tip bottom tube should be used to ensure fully washing of the precipitate.

      8. The protein content of the samples can’t be determined using the coomassie brilliant blue method.

      9. The protein content of the samples should be ranged from 1-10 mg/mL.

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      • Q&A (1)
      Q

      a********** Submitted [ Mar 02 2019 ]

      Asked: what is the normal rang value ? I got the value 0.353 . is this wrong ? and why ?

      Reply

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