Human Methemoglobin (MHB) ELISA Kit

The Human Methemoglobin (MHB) ELISA Kit is a high-sensitivity, sandwich ELISA designed for the quantitative detection of methemoglobin (MHB) in human serum, plasma, cell culture supernatants, tissue homogenates, and other biological fluids. MHB is a form of hemoglobin in which the iron is in the ferric (Fe³⁺) state and cannot bind oxygen. Elevated levels are associated with methemoglobinemia, toxic exposures, and oxidative stress. This kit enables researchers to measure MHB with high specificity and minimal sample volume, making it ideal for clinical and toxicological studies.

Key Features

• Assay Format: Sandwich ELISA
• Detection Range: 0.1 – 6 μg/mL
• Sensitivity: 0.01 μg/mL
• Sample Volume Required: 10 μL
• Detection Wavelength: 450 nm
• Storage Conditions: 2–8 °C
• Shelf Life: 6 months
• Precision:
  • Intra-assay CV <10%
  • Inter-assay CV <12%

Assay Principle

This kit uses a double-antibody sandwich ELISA approach. A microplate pre-coated with an anti-MHB antibody captures MHB from the sample. An HRP-conjugated detection antibody is then applied, followed by the addition of TMB substrate, which generates a colorimetric signal. The reaction is stopped with acid, turning the color from blue to yellow. The intensity of the signal is measured at 450 nm, and MHB concentration is calculated using a standard curve.

Kit Components

Validated Sample Types

• Serum / Plasma (EDTA, citrate)
• Cell Culture Supernatants
• Urine, CSF, and Pleural Fluids
• Tissue Homogenates

Note: Avoid sodium azide (NaN₃) and minimize freeze-thaw cycles. Samples can be stored at –20°C if not assayed immediately.

Assay Procedure Summary

1. Standard Dilution: Prepare 6-point dilution (9 → 0.5 μg/mL).
2. Add 40 μL sample diluent + 10 μL sample per well.
3. Incubate at 37 °C for 30 minutes, sealed.
4. Wash plate 5 times using 1× wash buffer.
5. Add 50 μL HRP-conjugate reagent to all wells (except blank), incubate 30 minutes.
6. Wash and add 50 μL each of Chromogen A and B; incubate 15 minutes in the dark.
7. Add 50 μL stop solution, read OD at 450 nm within 15 minutes.

Performance & Calculation

Data is analyzed using a log-log standard curve. Multiply the final concentration by the dilution factor (typically 5) for actual sample concentration.

The Human Methemoglobin (MHB) ELISA Kit (SL1179Hu) offers a reliable and sensitive method for detecting MHB in various human sample types. It is especially useful for studies investigating methemoglobinemia, oxidative stress, CO exposure, and drug-induced redox imbalances. With a low detection limit, small sample requirement, and robust intra-/inter-assay precision, this kit is an essential tool for translational toxicology and hematologic research.