MonkeyPox Virus IgM (MPV-IgM) ELISA Kit

The MonkeyPox Virus IgM (MPV-IgM) ELISA Kit is a qualitative enzyme-linked immunosorbent assay designed for the detection of Monkey MPV-specific IgM antibodies in serum, plasma, culture media, and other biological fluids.

This kit is optimized for research laboratories investigating early-stage immune responses to MonkeyPox Virus exposure. The assay employs antigen-coated microplate wells and an HRP–TMB colorimetric detection system to deliver reliable and reproducible results measured at 450 nm.

This product is strictly for research use and is not intended for therapeutic or diagnostic applications.

Intended Use

The kit is designed for qualitative detection of Monkey MPV-IgM antibodies in:

• Serum
• Plasma
• Culture media
• Urine and cerebrospinal fluid
• Cell culture supernatants
• Tissue homogenates

It is suitable for immunological and virology research applications involving MonkeyPox virus exposure and immune response studies.

Assay Principle

The assay is based on the ELISA technique:

1. Microplate wells are pre-coated with Monkey MPV-specific antigen.
2. IgM antibodies present in the sample bind to the immobilized antigen during incubation.
3. After washing, an HRP-labeled antigen conjugate binds to the captured antibodies, forming an antigen–antibody–enzyme complex.
4. Following a second wash, Tetramethylbenzidine (TMB) substrate is added, producing a blue color reaction.
5. The reaction is stopped with an acidic stop solution, resulting in a yellow endpoint color.
6. Optical density (OD) is measured at 450 nm using a microplate reader.
7. Results are interpreted by comparison with a calculated cut-off value.

The intensity of the color is proportional to the amount of MPV-IgM antibody present in the sample.

Kit Components

Each kit contains sufficient reagents for 48 or 96 determinations, including blank, negative, and positive controls.

Provided Components:

• Antigen-coated microplate
• Positive control
• Negative control
• HRP-conjugate reagent
• Sample diluent
• Chromogen Solution A
• Chromogen Solution B
• Stop solution
• Wash solution concentrate (20X for 48T; 30X for 96T)
• Adhesive films
• Instruction manual

Required Equipment (Not Included)

• ELISA microplate reader capable of measuring absorbance at 450 nm
• 37 °C incubator
• Plate washer (manual or automatic)
• Adjustable micropipettes (10–1000 µL range)
• Vortex mixer
• Distilled or deionized water
• Timer

Sample Preparation

Serum:

Allow blood to clot at room temperature for 10–20 minutes. Centrifuge at 2,000–3,000 rpm for 20 minutes and collect serum.

Plasma:

Collect blood in EDTA or citrate tubes. Centrifuge at 2,000–3,000 rpm for 20 minutes and collect plasma.

Urine and Other Fluids:

Centrifuge at 2,000–3,000 rpm for 20 minutes and collect supernatant.

Cell Samples:

Centrifuge culture supernatant and collect clarified fluid. For intracellular components, lyse cells in PBS (pH 7.2–7.4) using freeze–thaw cycles and centrifuge before testing.

Tissue Samples:

Homogenize tissue in PBS (pH 7.4) on ice, centrifuge at 2,000–3,000 rpm for 20 minutes, and collect supernatant.

Important Notes:

• Avoid repeated freeze–thaw cycles.
• Store samples at −20 °C if testing is delayed.
• Samples containing sodium azide (NaN₃) are not compatible due to inhibition of HRP activity.

Assay Workflow Summary

1. Add controls and diluted samples to designated wells.
2. Incubate at 37 °C for 30 minutes.
3. Wash plate five times with 1X wash buffer.
4. Add HRP-conjugate reagent and incubate at 37 °C for 30 minutes.
5. Wash plate five times.
6. Add Chromogen Solutions A and B and incubate at 37 °C for 15 minutes in the dark.
7. Add Stop Solution.
8. Measure absorbance at 450 nm within 15 minutes.

Thorough washing is critical to ensure assay precision and minimize background interference.

Result Interpretation

Validation Criteria:

• Average OD of Positive Control ≥ 1.00
• Average OD of Negative Control ≤ 0.10

Cut-Off Calculation:

Cut-Off = Average Negative Control + 0.15

Interpretation:

• OD < Cut-Off: Negative
• OD ≥ Cut-Off: Positive

Storage and Stability

• Store kit at 2–8 °C.
• Shelf life: 6 months.
• Unused plate strips should be resealed with desiccant and stored at 2–8 °C.
• Opened strips should be used within 1 month (maximum 3 months).

The MonkeyPox Virus IgM (MPV-IgM) ELISA Kit provides a standardized and efficient qualitative method for detecting early-phase MPV-specific IgM antibodies in a wide range of biological samples. Its antigen-coated microplate format, validated controls, and optimized HRP–TMB detection system ensure consistent and reproducible results for research laboratories engaged in virology and immunology studies.