製品の概要

  • 製品名

  • 検出方法

    Colorimetric/Fluorometric
  • サンプルの種類

    Cell culture supernatant, Urine, Other biological fluids, Tissue Extracts
  • アッセイタイプ

    Quantitative
  • 検出範囲

    0.01 nmol/well - 10 nmol/well
  • 全工程の試験時間

    0h 30m
  • 製品の概要

    Ascorbic Acid Assay Kit ab65346 provides a rapid, simple, and sensitive means of detecting ascorbic acid in various biological samples.


    In the ascorbic acid assay protocol, our proprietary catalyst oxidizes ascorbic acid to produce a product that interacts with the ascorbic acid probe, generating color and fluorescence. Ascorbic acid can be easily determined by either colorimetric (spectrophotometry at λ = 570 nm) or fluorometric (Ex/Em = 535/587 nm) methods.


    The assay can detect 0.01-10 nmol of ascorbic acid per assay.

  • 特記事項

    Ascorbic Acid (Vitamin C) plays an important role in many biological processes. It is a potent anti-oxidant, anti-inflammatory, anti-viral agent, and an immune stimulant and is present is a wide variety of foods and biological specimens. It is important to be able to monitor ascorbic acid content in these different samples.

  • 試験プラットフォーム

    Microplate reader

製品の特性

画像

  • Standard curve (fluorimetric) :  mean of duplicates (+/-SD) with background readings subtracted

  • Ascorbic Acid Standard Curve.

プロトコール

参考文献

ab65346 has not yet been referenced specifically in any publications.

レビューと Q&A

1-10 of 12 Abreviews or Q&A

Answer

I can confirm that the kit can be used with strawberry samples.

We recomend following guidelines for lysates preparation;

Homogenize the strawberries to homogeneity using the assay buffer in the kit and then centrifuge at top speed in a microcentrifuge to pellet debris. Collect the supernatant for the assay. It will be important to optimize the sample volume such that the solution the well does not become red since OD 570nm is in the pink

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Answer

Thank you for contacting us.

In this assay, our proprietary catalyst oxidizes ascorbic acid to produce a product that interacts with the ascorbic acid probe, generating color and fluorescence. So if the vitamin C is already oxidized, it is likely to generate color/fluorescence in the reaction without the catalyst. This value can then be subtracted from the +catalyst reaction readings for the same sample so that the amount of un-oxidized vitamin C can be calculated.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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Answer

Frozen samples should be thawed at room temperature, then kept on ice and used promptly in the assay (not warmed to room temperature). Ab65346 is recommended for your samples, since they are already diluted and will be further diluted during the assay. Ab65346 is more sensitive than ab65656 if using a fluorescent detection system.

If you don't have access to a fluorescent detection system, then either ab65346 or ab65656 will be fine to use.

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Question
Answer

Thank you for your inquiry.

The lab was able to provide this information:

1 mole/lt is 1 M.
This standard is 20 µmole. You reconstitute it in 200 µl. That is 20 µmole/200 µl (or 100 µmole/ml which is equivalent to 0.1 M or 100 mM.)
You are further diluting this 100 times. So it would be 20 µmole/20000 µl. That is equal to 20,000 nmole/20,000 µl. That is 1 nmol/µl. When you take 0,2,4,6,8 and 10 µl of this you are taking 0, 2,4,6,8 and 10 nmol/well of it.
I hope this information helps. Please contact us with any other questions.

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Question
Answer

Thank you for calling Abcam earlier today.

I have talked to the lab about whether the presence of EDTA in the samples would affect the kit and they say that it should be ok.

Please let me know if you have any more questions.

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Answer

I understand your concern. Signal from pre-existing carbonyl groups, before the catalytic generation of  carbonyl from ascorbic acid, would be an issue with other samples types too, including serum and plasma. The background contributed by sample proteins can be corrected for by substracting the signal produced by samples that do not receive the catalyst. I am hoping to receive some data illustrating this, which I will forward.

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Answer

Thanks for your call this week and for your patience while I have been in touch with the lab about ab65346. This kit does utilize a reaction on carbonyl groups, so it will likely cross-react with the PVP though this has not been specifically tested. As far as we can tell, you should be able to determine the amount of oxidized ascorbic acid by leaving out the catalyst, however this has also not been tested. I hope this information will be useful in determining whether this kit is appropriate for your studies, but please let me know if you have any further questions.

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Answer

Thank you for your inquiry. I just heard back from the testing laboratory that these kits cannot distinguish ascorbic acid isoforms. I am sorry that this information could not be more helpful, but please let us know if we can be of any other help.

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Answer

Thank you for contacting us. I have hear back from the ab with the following information to your questions: 1. Yes, you can use plasma samples – but for ab65656 we recommend removing the proteins (e.g. by using spin filter (10 kDa cutoff) (ab93349) or the Deproteinizing Sample Preparation Kit (ab93299)). 2. The proteins can be removed after thawing your plasma samples. 3. EDTA in your plasma samples is probably OK, but I am still waiting for a definite answer from the lab. I will let you know when I have the information. 4. The kit ab65656 is the best to use with your samples. 5. Yes, the power of this assay kit ab65656 is that it determines ONLY ascorbate by eliminating (subtracting) contributions for any other anti-oxidants. To do this need a background sample for EACH test sample. That means you would need three background samples for the 3 samples per patient. I hope this information is so far helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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Answer

Thank you for your inquiry. Of the two kits, ab65656 is more suitable. Theoretically ab65656 should be compatible with plant samples as long as the lysates are prepared in the assay buffer provided in the kit. Technically though, since we have not tested this kit for compatibility with plant samples, this has to be optimized and we can not guarantee that it will work with plant samples. The plant samples can be homogenized on ice in 2-3 volumes of the assay buffer using a Dounce homogenizer/ mortar-pestle. This has to be centrifuged to remove the cell debris, and the supernatant can then be used for the assay. I hope this information helps. Please contact us with any other questions.

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