Description
Hydrogen Peroxide Assay Kit (BA0047) (BA0047)
The Hydrogen Peroxide Assay Kit (SKU: BA0047) provides a simple, direct and automation-ready method for the quantitative colorimetric determination of peroxide in biological samples without any pretreatment. Peroxide, such as hydrogen peroxide, is one of the key reactive oxygen species formed under oxidative stress, and high levels have been linked to pathological conditions including ageing, asthma, diabetes, atherosclerosis, cataract, inflammatory arthritis and neurodegenerative diseases. The kit uses the chromogenic Fe3+-xylenol orange reaction, in which a purple complex forms when Fe2+ in the reagent is oxidised to Fe3+ by peroxides present in the sample. The intensity of the colour, measured at 540-610nm, is an accurate measure of the peroxide level. Enhanced colour intensity is achieved using sorbitol, and the optimised formulation substantially reduces interference by substances in raw samples.
| Product Name: | Hydrogen Peroxide Assay Kit (BA0047) |
| SKU: | BA0047 |
| Detection Method: | Colorimetric |
| Detection Range: | 0.2 µM (7 ng/mL) to 30 µM (1,020 ng/mL) H2O2 in 96-well plate assay |
| Sample Type: | Serum, citrate-plasma, urine, cell lysate, culture medium |
| Species Reactivity: | All |
| Assay Time: | 30 minutes |
| Kit Size: | 250 Assays |
| Equipment Required: | Microplate reader |
| Storage: | The kit is shipped at room temperature. Store all reagents at 4°C. |
| Shelf Life: | 12 months after receipt |
| Shipping: | Room Temperature |
This kit measures peroxide concentration in biological samples without pretreatment. The improved method utilises the chromogenic Fe3+-xylenol orange reaction, in which a purple complex forms when Fe2+ provided in the reagent is oxidised to Fe3+ by peroxides in the sample. The intensity of the colour, measured at 540-610nm, is an accurate measure of the peroxide level.
- Sensitive and accurate: enhanced colour intensity using sorbitol; detection range 0.2 µM (7 ng/mL) to 30 µM (1,020 ng/mL) H2O2 in 96-well plate assay
- Simple and high-throughput: addition of a single detection reagent and incubation for 30 minutes, readily automated for thousands of samples per day
- Direct assays: H2O2 in biological samples (e.g. serum, citrate-plasma, urine, cell lysate, culture medium)
- Pharmacology: effects of drugs on peroxide metabolism
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 | Prepare Detection Reagent by mixing 1 volume Reagent A with 100 volumes Reagent B; equilibrate to room temperature before assay. |
| 2 | Standards: prepare fresh standards on the day of assay. Pipette 5 µL 3% H2O2 and mix with 495 µL H2O, then mix 5 µL of this solution with 1465 µL H2O to give a 30 µM Premix. Dilute standard as shown in the table. |
| 3 | Transfer 40 µL diluted standards and each sample into separate wells of a clear flat-bottom 96-well plate. Add 200 µL Detection Reagent to all standards and samples. |
| 4 | Incubate 30 minutes at room temperature and read optical density at 540-610nm (peak absorbance at 585nm). |
Subtract blank OD (water, #8) from the standard OD values and plot OD against H2O2 concentrations. Subtract blank OD from Sample OD and determine the sample peroxide content from the standard curve. Conversion: 1 µM H2O2 equals 34 ng/mL or 34 ppb.
| Component | Quantity | Storage |
| Reagent A | 1 mL | 4°C |
| Reagent B | 50 mL | 4°C |
| Standard (3% stabilised H2O2) | 100 µL | 4°C |
