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    Quest 5-hmC DNA ELISA Kit


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    Cat # Name Size
    D5425 Quest 5-hmC DNA ELISA Kit 1 X 96 Preps
    D5426 Quest 5-hmC DNA ELISA Kit 2 X 96 Preps

    Highlights

    • Sensitive and specific detection of 5-hydroxymethlycytosine (5-hmC) from a variety of samples
    • Ideal for global 5-hmC quantitation and high throughput screening
    • Streamlined workflow can be completed in 3 hrs
    Description & Documents Specifications FAQ Components Citations

    Documents

    Product Description

    The Quest 5-hmC DNA ELISA Kit is both sensitive and specific and can be used to accurately detect 5-hmC DNA in a variety of samples. The kit is compatible with a wide range of input DNA including intact vertebrate, plant, and microbial genomic DNA, as well as enzyme-digested and mechanically sheared fragments. The Control DNA Set included with this kit has been calibrated to accurately quantify the percent 5-hmC in sample DNA by use of a standard curve. Also, the fast, streamlined workflow is ideal when analyzing/screening large numbers of samples.

    Documents

    Technical Specifications

    Detection ≥ 0.02% 5-hmC per 100 ng DNA
    Equipment Incubator and ELISA plate reader. A multi-channel pipettor is recommended. An automated plate washer may be used for blocking and wash steps.
    Input This protocol is optimized for 100 ng of input DNA/well. Compatible with DNA in the range of 20-200 ng/well.
    Sample Source Purified genomic DNA in water, Tris-EDTA, or similar.

    Resources

    We guarantee integrity of kit components for up to 6 months from date of purchase

    Shipping of components is on blue and dry ice. Upon arrival, make sure to store buffers at 4°C, Control DNA and Anti-5hmC antibody at -20°C, and the Anti-DNA antibody can be stored at -20°C for 1 week (For long-term storage keep at -80°C).

    The DNA ELISA controls are genomic DNA. Control A is E. coli genomic DNA as it comprises 0% 5-hmC. The other controls derive from various mouse tissues and the %5-hmC is specific to each tissue. The levels of 5-hmC are were quantify by mass-spec.

    The optimal absorbance is at 405 nm, however 405-450 nm will work as well.

    To calculate global %5-hmC, control DNAs and samples must be assayed on the same plate. Use the equation x=y-b/m and solve for x (%5-hmC).

    1. Make sure the correct buffers were used to coat, block and wash the plate. 2. Double-check that HRP developer and the Anti-DNA HRP antibody were stored and prepared correctly.

    1. Make sure that standard curve developed properly (R2 > 0.9). 2. Verify that all samples and controls are prepared correctly, mixed well before dilution and addition to the wells. 3. Ensure that anti-DNA HRP antibody was stored and prepared correctly before addition to the wells. 4. %5-hmC levels of your sample might be too low for the assay to detect (detection limit is 0.01% in 100 ng of denatured DNA).

    Cat # Name Size
    A4001-25 Anti-5-Hydroxymethylcytosine Polyclonal Antibody 25 µg
    D5425-5-1 Control A (100 ng/µl) 40 µl
    D5425-5-2 Control B (100 ng/µl) 40 µl
    D5425-5-3 Control C (100 ng/µl) 40 µl
    D5425-5-4 Control D (100 ng/µl) 40 µl
    D5425-5-5 Control E (100 ng/µl) 40 µl
    D5425-4-15 HRP Developer 15 ml
    D5425-1-15 Coating Buffer 15 ml
    D5425-2-30 10X ELISA Buffer 30 ml
    D5425-3-100 Anti-DNA HRP Antibody (100X) 100 µl
    C2020 96-Well ELISA Plate 12 x 8-Well Strips

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