A nitrocellulose membrane based solid phase enzyme immunoassay for detecting polioviruses in stool.

Area of Science:

• Virology
• Immunology
• Biotechnology

Background:

• Enzyme immunoassays are crucial for detecting viral pathogens.
• Nitrocellulose membranes offer potential as solid-phase supports in immunoassays.
• Optimizing blocking agents and sample preparation is key for assay sensitivity and specificity.

Purpose of the Study:

• To develop and optimize an enzyme immunoassay for poliovirus detection.
• To evaluate nitrocellulose membrane as a solid-phase support.
• To identify critical factors influencing assay performance, including blocking agents and sample treatment.

Main Methods:

• Development of a 96-well enzyme immunoassay using nitrocellulose membrane.
• Comparison of monoethanolamine and skimmed milk as blocking agents.
• Assessment of stool specimen collection medium (skimmed milk percentage), dilution buffer composition/pH, and chloroform treatment.
• Evaluation of polyvinyl affinity membrane versus nitrocellulose membrane.

Main Results:

• The developed immunoassay detected up to 10(4.28) TCID50 poliovirus particles/0.1 ml of stool.
• Monoethanolamine demonstrated superior performance as a blocking agent compared to skimmed milk.
• Key factors influencing specificity included skimmed milk concentration, buffer properties, and pre-treatment of stool specimens.
• Nitrocellulose membrane performed comparably to polyvinyl affinity membrane.

Conclusions:

• An effective enzyme immunoassay for poliovirus detection in stool was established using nitrocellulose membrane.
• Monoethanolamine is a recommended blocking agent for this assay, outperforming skimmed milk.
• Careful optimization of sample collection and processing is essential for reliable poliovirus detection.