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Iron Assay Kit (Colorimetric) (ab83366)

Reviews (3)Q&A (19)References (234)
Functional studies - ab83366
  • Functional Studies - Iron Assay Kit (ab83366)
  • Functional Studies - Iron Assay Kit (ab83366)
  • Functional Studies - Iron Assay Kit (ab83366)
  • Functional Studies - Iron Assay Kit (ab83366)

Key features and details

  • Assay type: Quantitative
  • Detection method: Colorimetric
  • Platform: Microplate reader
  • Assay time: 1 hr 30 min
  • Sample type: Other biological fluids, Serum, Urine
  • Sensitivity: 8 µM

製品の概要

  • 製品名

    Iron Assay Kit (Colorimetric)
    Iron キット 製品一覧

  • 検出方法

    Colorimetric

  • サンプルの種類

    Urine, Serum, Other biological fluids

  • アッセイタイプ

    Quantitative

  • 検出感度

    > 8 µM

  • 検出範囲

    8 µM - 400 µM

  • 全工程の試験時間

    1h 30m

  • 製品の概要

    Iron Assay Kit ab83366 provides a simple convenient means of measuring ferrous (Fe2+) and/or ferric (Fe3+) iron in biological samples.


    Ferric carrier protein will dissociate ferric into solution in the presence of the acid assay buffer. If the assay buffer has a neutral pH, iron will bind tightly to the iron carrier protein. However, under acidic conditions (pH less than 5.5), iron no longer has a binding affinity for the carrier protein and will be dissociated and released into the solution as iron/ferric ions, whose concentration you can measure using this kit. The Iron Assay Buffer in this kit has an acidic pH that enables this release of iron/ferric ions into the solution.


    Free ferrous iron (Fe2+) reacts with Iron Probe to produce a stable colored complex with absorbance at 593 nm. Ferric iron (Fe3+) can be reduced to form Fe2+ enabling the measurement of total iron (Fe2+ and Fe3+). The level of ferric iron (Fe3+) is calculated by subtracting ferrous iron from total iron.


    A specific chelate chemical is included in the buffer to block copper ion (Cu2+) interference.
    The kit measures iron in the linear range of 0.4 to 10 nmol or 8 µM to 200 µM.


    Iron assay protocol summary:
    - add samples and standards to wells
    - for Fe2+ assay add assay buffer only, for total iron (Fe2+ and Fe3+) add iron reducer
    - incubate for 30 min at 37ºC
    - add Iron Probe and incubate for 60 min at 37ºC
    - analyze with microplate reader

  • 特記事項

    This product is manufactured by BioVision, an Abcam company and was previously called K390 Iron Colorimetric Assay Kit. K390-100 is the same size as the 100 test size of ab83366.

  • 試験プラットフォーム

    Microplate reader

製品の特性

画像

  • Functional studies - ab83366
    Functional studies - ab83366Leal SM Jr et al., PLoS Pathog 9(7), fig 2c. doi: 10.1371/journal.ppat.1003436. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

    Leal SM Jr et al examined if iron availability regulates fungal growth in an infection as fungal infection intiates an iron sequestration response. Mice given Fe-dextran (Fe-Dex) and deferoxamine (Defox) shows a higher fungal mass (Fungal dsRed) compared to vehicle treated mice over 48 hours. Iron content was quantified in mouse serum using Iron assay kit (ab83366).

  • Functional Studies - Iron Assay Kit (ab83366)
    Functional Studies - Iron Assay Kit (ab83366)

    Iron measured in human urine showing concentration (micromolar)

  • Functional Studies - Iron Assay Kit (ab83366)
    Functional Studies - Iron Assay Kit (ab83366)

    Assay of soluble free iron from a soil sample (5 μL of 100 μL buffer into which 100 mg of soil had been stirred), 5 μL of FBS and 5 μL of a 100 μM sample of iron standard.

  • Functional Studies - Iron Assay Kit (ab83366)
    Functional Studies - Iron Assay Kit (ab83366)

    Example of iron standard curve using ab83366.

  • Functional Studies - Iron Assay Kit (ab83366)
    Functional Studies - Iron Assay Kit (ab83366)

    Assay of total iron(II + III) in off-the-clot human serum (50 µl of serum was added per well). Data are mean ± SEM from 2 independent replicates, performed in duplicate wells.

参考文献 (234)

ab83366 を使用した論文を発表された方は、こちらまでお知らせください。データシートに掲載させていただきます。

ab83366 は 234 報の論文で使用されています。

  • Xiong L  et al. Downregulation of TNFAIP1 alleviates OGD/R‑induced neuronal damage by suppressing Nrf2/GPX4‑mediated ferroptosis. Exp Ther Med 25:25 (2023). PubMed: 36561622
  • Zhang K  et al. Heat shock protein 27 deficiency promotes ferrous ion absorption and enhances acyl-Coenzyme A synthetase long-chain family member 4 stability to promote glioblastoma cell ferroptosis. Cancer Cell Int 23:5 (2023). PubMed: 36639654
  • Pitts HA  et al. SPINK2 Protein Expression Is an Independent Adverse Prognostic Marker in AML and Is Potentially Implicated in the Regulation of Ferroptosis and Immune Response. Int J Mol Sci 24:N/A (2023). PubMed: 37298647
  • Daoud L  et al. Elucidating the effect of iron acquisition systems in Klebsiella pneumoniae on susceptibility to the novel siderophore-cephalosporin cefiderocol. PLoS One 17:e0277946 (2022). PubMed: 36580460
  • Ma X  et al. Lidocaine attenuates hypoxia/reoxygenation-induced inflammation, apoptosis and ferroptosis in lung epithelial cells by regulating the p38 MAPK pathway. Mol Med Rep 25:N/A (2022). PubMed: 35244190
View all Publications for this product

レビューと Q&A

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レビューを送る 質問を送る

1-10 of 22 Abreviews or Q&A

Using extract from murine colon feces for iron determination.

 Excellent Excellent 5/5 (Ease of Use)
Abreviews
abreview image
I used pools of sera and colon poo extract to assess the utility of the kit for different samples. These samples were from Apodemus sylvaticus mice.
Sample preparation:
1. Sera was defrosted after storage at -80C. Centrifuged defrosted sera at 12000 rpm for 4mins.
2. Avoided disrupting the pellet and pipetted off equal volumes of sera into a clean Eppendorf to form a pool.
3. Colon poo was also defrosted from -80C. I weighed the sample and multiplied this mass by 3, and added this volume of proteinase inhibitor solution (PIS). For 0.2g of colon poo, a volume of 600ul PIS was added.
a. PIS was prepared by dissolving one tablet of Complete Mini Protease Inhibitor Cocktail Tablets (Roche, 11836153001) in 10ml 1x PBS.
4. Vortexed poo and PIS for 5 minutes. Stood for an hour at room temperature. Centrifuged at 12000rpm for 5 minutes. Pipetted equal volumes of supernatant into a clean Eppendorf to form the faecal extract pool.
5. Make a 1mM standard by diluting 2ul standard into 198ul Dh20.
6. Diluted samples and standards into wells with assay buffer.
Dilutions are listed.
a. Standard, in final volumes of 50ul and 100ul. 2nmol/well, 4nmol/well, 6nmol/well, 8nmol/well, 10nmol/well.
b. Sera 50ul volume, 1:1, 2:5, 1:5, 1:50.
c. Sera 100ul volume 1:2, 1:5, 1:10, 1:100.
d. Fecal extract 50ul volume, 1:1, 2:5, 1:5, 1:50, 1:100.
e. Fecal extract 25ul volume 1:1, 1:2, 1:5, 1:10, 1:100.
7. Add 5ul iron reducer to 100ul samples and standards. Add 2.5ul to 50ul samples.
8. Add 100ul probe to 100ul wells and 50ul probe to 50ul wells.
9. Incubate for 1hr protected from light at 37 degrees C.
10. Read at 593nm.

Conclusions:
I was testing whether for a small volume of sample it is better to dilute the sample into a large volume or to have the sample at a higher concentration in a lower volume. Both volumes worked equally well here. The poo pool had lower readings than the serum pool but was still detectable. Serum volumes of 5-10ul or faecal extract volume of 20ul would be suitable for future use. I used the two standard curves of different volumes to calculate concentrations for each pool. I averaged readings across dilutions. The concentration of the pools was 87uM for faecal extract in 105uL total well volume or 89uM in 205uL well volume. Sera was 205uM in 105uL total well volume or 219uM in 200uL. Both protocols gave congruent readings.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

投稿 Dec 20 2022

Iron assay Kit ab 83366

 Excellent Excellent 5/5 (Ease of Use)
Abreviews
The product is working with satisfactory reading using Serum and tissue lysate. In the next step i am testing in rabbit CSF for Iron forms II and III.
I will update/ upload data sets once i complete CSF analysis.
The kit is working total iron assay for rabbit templates ( Serum and Lysate)
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

投稿 Feb 09 2022

Detection of iron levels in hair samples

 Average Good 4/5 (Ease of Use)
Abreviews
We used the iron assay kit to evaluate whether iron can be detected in hair shaft samples. In this case, we used a hair sample from a Rhesus primate, since we had a sufficient amount to test. The hair sample was milled into a fine powder, homogenized with assay buffer (4x volume) and centrifuged as directed. We initially tested various small amounts of hair (ranging from 1.8 to 12.1mg), given that these samples are often only available in limited quantities. The resulting absorbance was very close to the base nm of the 0 standard, and translated to -0.043 to 0.034nmoles per 50μl sample. We had used two sets of standard concentrations: the set of standards recommended in the protocol (0-10nmol/well) and a 1:10 dilution of those standards (0-1nmol/well). Both standard curves had an R-value better than 0.98, and the diluted standard curve was used since it best encompassed the resulting nm.
We repeated the assay with a larger amount of hair (55.9mg). In the increased sample we were able to detect 0.14-0.23nmole per 50μl sample, which was closer to the stated range of the kit (0.4-20 nmole/50μl). This indicates that it is possible to detect iron in hair shaft sample using this kit. However, given the relatively large amount of sample needed, it is not an ideal method for limited sample sizes. The kit itself was very easy to use.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

DR. Erilynn Heinrichsen

Verified customer

投稿 Nov 05 2013

Question

Can I use this kit with plasma samples?

Read More
Answer

Unfortunately, this kit is not suitable for use with plasma samples due to the presence of iron binding transferrin in plasma leading to inaccurate measurements of free iron.

Read More

Question

My plate reader reads at 570 nm, but I see this kit requires 593 nm. Could you send me the emission spectra for the probe, so I can determine whether or not my platereader would be compatible?

Read More
Answer

Since the specific probe is proprietary we do not have a spectrum to share, but in our experience reading at 570 nm should work and may simply reduce the sensitivity slightly.

The emission profile shows a sharp peak at 580-590 nm, can measure signal within 590 +/- 20 nm.

Read More

Question

i want to measure ferrous and ferric acid separately, can I do that with this kit?

Read More
Answer

Yes, you can definitely measure ferrous and ferric acid separately with this kit. Please follow the protocol for measuring the ferrous and the total iron in the samples. When you subtract the ferrous levels from the total level, you get the ferric levels in the samples.

Read More

Question

During sample prep we are seeing cloudiness form but not blue color when using mouse liver samples. Samples are homogenized in Assay buffer with dounce homogenizer and spun at 16,000 x g as recommended. We have used straight attempted dilution of the samples 1/10 & 1/100. The kit is new and has been stored correctly.

Read More
Answer

This is a common problem seen in liver and serum samples. Lipoproteins in the sample are the main culprits behind this turbidity.
For this purpose, we would recommend that to add 5 µl/well of 1 M SDS (28.8% or 288 mg/ml of SDS) to all their sample wells after step 4.2. Incubate for 30 min at 25 °C as mentioned in the datasheet and then follow the protocol. This SDS will clear up the turbidity (by dissolving any lipoproteins in the samples).

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Question

Can I use samples that have been prepared using RIPA buffer?

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Answer

We do not typically recommend using RIPA buffer with our kits since it contains SDS which can denature proteins but for this assay it could work. We have not tested with RIPA buffer. We recommend using the assay buffer that comes with the kit.

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Question

Does the assay also measure heme bound iron? or just the so called free iron.

Read More
Answer

I can confrim that the ab83366 Iron Assay Kit will measure the free ferrous and ferric forms of iron and not the iron complexed in Heme.

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Question

I have a couple questions about this kit:
1. How are the reagents to be stored between uses?
2. Does the plate need to be covered during incubations?

Read More
Answer

1. All components should be stored in aliquots according to individual use requirements at -20C to prevent freeze-thaw cycles.

2. The plate can be covered by a plate cover and aluminium foil to protect from light during incubation. The incubation is at RT and hence there is not much concern about evaporation at 25C.

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1-10 of 22 Abreviews or Q&A

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