ELISA Kits & Reagents

<h2>What are 5-hmC ELISA kits?</h2>
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5-hmC ELISA kits are antibody-based assays formulated specifically to quantify 5-hydroxymethylcytosine, the oxidized derivative of 5-methylcytosine produced by TET family enzymes, in a genomic DNA sample. The assay typically uses denatured single-stranded DNA coated on a plate well, recognized by an anti-5-hmC primary antibody and developed with an HRP-conjugated secondary antibody for a colorimetric readout proportional to the global level of 5-hmC in the sample.
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<h2 dir="ltr">What you will find:</h2>
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<ul>
<li dir="ltr" role="presentation">DNA hydroxymethylation ELISA assay kits for 5-hmC levels quantification</li>
<li dir="ltr" role="presentation">DNA methylation ELISA kits for comparative gene regulation analysis and measurement of 5-mC&nbsp;</li>
<li dir="ltr" role="presentation">DNA standards for accurate methylation and hydroxymethylation quantification&nbsp;</li>
<li dir="ltr" role="presentation">Control DNA sets and assay controls for experimental accuracy, validation, and reproducibility</li>
<li dir="ltr" role="presentation">HRP-based antibodies for the specific detection in ELISA workflows</li>
<li dir="ltr" role="presentation">ELISA coating and reaction buffers for enhanced assay performance and setup</li>
<li dir="ltr" role="presentation">Signal development reagents for quantitative readout and chemiluminescent or colorimetric detection</li>
</ul>
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<h2>How to use 5-hmC ELISA kits</h2>
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Confirm 5-hmC is detectable at relevant levels in your cell type or tissue
5-hmC is most abundant in neurons and embryonic stem cells, where it can constitute up to 20% of total modified cytosines in neuronal DNA, while most cultured cell lines have much lower 5-hmC levels that may fall below a plate-based antibody assay's reliable detection limit. Checking published 5-hmC abundance for your specific cell type helps confirm whether ELISA-level detection is appropriate before committing samples to the assay.
Use a set of mixed methylation standards to build a calibration curve
Running defined mixtures of 5-hmC-containing and unmethylated DNA at known ratios generates a standard curve that lets you convert the raw absorbance signal to an actual global hydroxymethylation percentage, since the assay signal alone cannot be interpreted as an absolute level without this calibration.
Confirm your antibody's specificity for 5-hmC versus 5-mC
Anti-5-hmC antibodies used in these kits are selected for specificity to 5-hydroxymethylcytosine rather than 5-methylcytosine, but published cross-reactivity data for the specific antibody in your kit is worth checking, particularly when studying sample types where both marks are present at similar levels.
Plan your experimental design to include appropriate negative and positive controls
Including a negative control (unmethylated DNA, expected to give background-level signal) and a positive control (5-hmC-containing DNA of known level) in every plate confirms the assay is performing as expected and catches plate-to-plate variability before it affects result interpretation.
Understand ELISA gives global level, not locus-specific 5-hmC information
A 5-hmC ELISA measures the average hydroxymethylation level across all DNA in the sample; it does not reveal which specific CpG sites carry 5-hmC, which requires hMeDIP-seq or TAB-seq for regional or base-level resolution respectively.
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<h2>Specifications context</h2>
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Anti-5-hmC antibodies used in ELISA formats are validated specifically for 5-hydroxymethylcytosine and selected for minimal cross-reactivity with 5-methylcytosine, since distinguishing these two modifications is the primary purpose of a dedicated 5-hmC assay rather than a general methylation assay. As of 2026, 5-hmC ELISA kits remain a standard first-pass tool in studies where many samples need to be screened for differences in global hydroxymethylation level before committing a smaller subset to high-resolution sequencing methods.
<a href="https://mbpinc.net/contact-us">Contact the expert team</a> at Molecular Biology Products Inc. (MBP) and find reliable ELISA kits and reagents for your lab.

MSDS

10X ELISA Buffer

For use with Zymo Research purification products.

Manufacturer

Shipping Condition

Storage Condition

10X ELISA Buffer (30 ml)

10X ELISA Buffer (60 ml)

5-Hydroxymethylcytosine DNA Standard (2 µg)

D5405-3 by Zymo Research is a 5-Hydroxymethylcytosine DNA Standard — a 897 bp linear dsDNA in which 100% of cytosines are 5-hmC. Each vial contains 2 µg at 50 ng/µl for use as a positive control or calibration reference in 5-hmC ELISA, dot blot, and LC-MS/MS workflows.

Concentration

Sample Storage

Protocol

5-Methylcytosine & 5-Hydroxymethylcytosine DNA Standard Set

The D5405 DNA Standard Set by Zymo Research contains three sequence-identical 897 bp dsDNA standards at 50 ng/µl (2 µg each): unmodified cytosine (D5405-1), 5-methylcytosine (D5405-2), and 5-hydroxymethylcytosine (D5405-3). Ideal reference controls for HPLC, LC/MS, and TLC cytosine modification quantitation studies.

Applicable For

5-Methylcytosine DNA Standard (2 µg)

5-Methylcytosine DNA Standard (2 µg) (D5405-2) by Zymo Research — fully synthetic 897 bp dsDNA with 100% 5-mC at all cytosines. 50 ng/µl, 40 µl (2 µg total) in modified TE buffer. For bisulfite conversion controls, MeDIP, LC/MS, and HPLC assays.

Anti-DNA HRP Antibody (100X)

For use with Zymo Research ELISA products

Anti-DNA HRP Antibody (100X) (100 µl)

Anti-DNA HRP Antibody (100X) (200 µl)

Coating Buffer

D5425-1-15 by Zymo Research — Coating Buffer, 15 mL; component of the Quest 5-hmC DNA ELISA Kit; used in the microplate coating step for 5-hydroxymethylcytosine (5-hmC) detection and quantification. For research use only.

Coating Buffer (15 ml)

Coating Buffer (30 ml)

Control A (100 ng/ul) (40 µl)

D5425-5-1 by Zymo Research is Control A, a 40 µl aliquot of 100 ng/µl control DNA for use with the Quest 5-hmC DNA ELISA Kit for 5-hydroxymethylcytosine detection and quantification. Store at -20°C. Research use only.

Control B (100 ng/ul) (40 µl)

D5425-5-2 by Zymo Research is Control B, a calibrated mouse genomic DNA standard at 100 ng/µl (40 µl volume) for use as a reference point in the Quest 5-hmC DNA ELISA Kit (D5425/D5426) standard curve. Used to quantify 5-hydroxymethylcytosine (5-hmC) in unknown samples at ≥0.02% sensitivity per 100 ng DNA. Must be run on the same ELISA plate as unknowns. Store at −20 °C.

Control C (100 ng/ul) (40 µl)

D5425-5-3 by Zymo Research is Control C, a mouse tissue-derived genomic DNA at 100&nbsp;ng/&micro;l (40&nbsp;&micro;l) with defined %5-hmC for standard curve construction in the Quest 5-hmC DNA ELISA Kit (D5425). Research use only.

Control D (100 ng/ul) (40 µl)

D5425-5-4 Control D by Zymo Research — 100 ng/µl, 40 µl, 0.23% 5-hmC — intermediate calibration standard for the Quest 5-hmC DNA ELISA Kit standard curve. Store at -20°C.

Control DNA Set (5 x 40 µl)

D5425-5-C by Zymo Research is the Control DNA Set (5 × 40 µl), providing calibration standard DNA for use with the Quest 5-hmC DNA ELISA Kit to detect and quantify 5-hydroxymethylcytosine (5-hmC) in genomic DNA. Required for generating the ELISA calibration curve. For research use only.

Control E (100 ng/ul) (40 µl)

D5425-5-5 by Zymo Research is Control E, a mouse tissue-derived genomic DNA at 100&nbsp;ng/&micro;l (40&nbsp;&micro;l) with defined %5-hmC for standard curve construction in the Quest 5-hmC DNA ELISA Kit (D5425). Used alongside Control C (D5425-5-3). Research use only.