Description
DNA Assay Kit (Fluorometric) (BA0171) (BA0171)
The DNA Assay Kit (Fluorometric) (SKU: BA0171) is designed to accurately measure nanogram quantities of double-stranded DNA where the traditional UV 260 nm absorbance method lacks sensitivity. Accurate determination of DNA concentration, particularly at low concentrations, is crucial for reproducible results in sequencing, cloning, transfection and DNA labelling. The improved method utilises a Hoechst dye that binds specifically to double-stranded DNA, and the resulting fluorescence, measured at 450 nm (lambda exc = 350 nm), is directly proportional to the DNA concentration in the sample. The optimised formulation substantially reduces interference from substances present in raw samples, tolerating RNA, salt up to 3 M NaCl, detergent below 0.01% SDS and common DNA extraction buffers. The simple mix-and-read procedure can be readily automated for high-throughput screening. It is intended for research use only.
| Product Name: | DNA Assay Kit (Fluorometric) (BA0171) |
| SKU: | BA0171 |
| Detection Method: | Fluorimetric (lambda ex/em = 350/450 nm) |
| Detection Range: | 2 ng to 40 ng (100-2,000 ng/mL) calf thymus DNA in 96-well plate assay |
| Sample Type: | Plasmid DNA, genomic DNA, cDNA, PCR products and DNA extracted from gels |
| Species Reactivity: | All |
| Assay Time: | Approximately 1 min incubation |
| Kit Size: | 250 Assays |
| Equipment Required: | Microplate reader |
| Storage: | DNA standard at -20C; Reagent at 2-8C |
| Shelf Life: | 12 months after receipt |
| Shipping: | Room Temperature |
A sensitive fluorimetric assay for the quantitation of nanogram quantities of double-stranded DNA. The method uses a Hoechst dye that binds specifically to dsDNA, producing a fluorescence signal at 450 nm (lambda exc = 350 nm) that is directly proportional to DNA concentration. The linear detection range is 2-40 ng (100-2,000 ng/mL) calf thymus DNA in a 96-well plate assay, with low interference from RNA, salt, detergent and common extraction buffers.
- Sensitive and accurate; linear detection range 2 ng to 40 ng (100-2,000 ng/mL) calf thymus DNA
- Simple mix-and-read procedure using a single working reagent
- Readily automated for high-throughput measurement of thousands of samples per day
- Low interference from RNA, salt (up to 3 M NaCl), detergent (< 0.01% SDS) and common extraction buffers
- Direct assay of plasmid DNA, genomic DNA and cDNA
- Quantitation of DNA following polymerase chain reaction
- Measurement of DNA extracted from gels and other matrices
Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
| Step | Procedure |
| 1 | Bring reagents to room temperature before use. Prepare 400 uL of 2000 ng/mL Premix by mixing 80 uL Standard and 320 uL TE buffer (10 mM Tris, 1 mM EDTA, pH 7.4), then dilute the standards as shown in the table. |
| 2 | Transfer 20 uL of diluted standards and samples into wells of a black flat-bottom 96-well plate (store standards at 4C for future use). |
| 3 | Add 200 uL working reagent and tap lightly to mix. |
| 4 | Incubate for 1 minute at room temperature and read fluorescence emission at 440-460 nm (peak 450 nm, excitation 340-370 nm). |
| 5 | Cuvette procedure: transfer 100 uL diluted standards and samples to cuvettes, add 1000 uL working reagent, tap to mix, incubate 1 minute and read at 440-460 nm. |
Subtract the blank fluorescence value (water, #8) from the standard values and plot delta F against standard DNA concentrations. Determine the slope by linear regression. [DNA] = (F_SAMPLE - F_BLANK) / Slope x n (ng/mL), where F_SAMPLE and F_BLANK are the fluorescence values of the sample and the water or buffer in which the sample was diluted, respectively, and n is the sample dilution factor.
| Component | Quantity | Storage |
| Reagent | 50 mL | 2-8C |
| Standard (10 ug/mL calf thymus DNA) | 1 mL | -20C |
