Anti Phocaeicola, Monoclonal Antibody (PV-L2B7-117K1)

Developed by Dr. Jun Kunisawa and Dr. Ken Yoshii at the National Institutes of Biomedical Innovation, Health and Nutrition, Fujifilm Wako now offers monoclonal antibody against Phocaeicola vulgatus¹⁾ (formerly Bacteroides vulgatus). It is suitable for use in ELISA, flow cytometry, immunoprecipitation, and Western blotting, allowing for rapid, simple, and cost-effective detection of gut bacteria.

Antibody Information

Quantification of Individual Gut Bacterial Species by Sandwich ELISA and Comparison with 16S rRNA Gene-Based NGS

Bacterial counts in three human fecal samples were measured using sandwich ELISA with combinations of anti-gut bacterial antibodies. Relative abundances obtained by sandwich ELISA were compared with those obtained from 16S rRNA gene amplicon sequencing via NGS¹⁾.

Quantification of Gut Bacterial Counts by Sandwich ELISA

• Phocaeicola vulgatus
• Faecalibacterium duncaniae
• Segatella copri

Comparison with 16S rRNA Gene-Based NGS

• Phocaeicola vulgatus
• Faecalibacterium duncaniae
• Segatella copri

ELISA

100 μL of a bacterial suspension was added to a 96-well microplate to coat the wells. The suspension was prepared either by diluting freeze-dried, heat-inactivated bacteria in PBS or by suspending them in B-PER solution followed by lysis with glass beads. After blocking and washing the antigen-coated plate, culture supernatants from individual hybridoma clones were added and incubated for 2 hours. Antibodies against gut bacteria bound to the antigens were detected using HRP-conjugated goat anti-mouse IgG¹⁾.

• P. vulgatus
• F. duncaniae
• S. copri
• B. pseudocatenulatum
• B. longum
• B. wexlerae
• A. muciniphila

Flow Cytometry

After fixation with Farmer’s fixative (ethanol:acetic acid = 7:3), the bacteria were centrifuged at 13,040 x g for 2 minutes at 4 °C. Next, 100 μL of the bacterial suspension was mixed with 1 μg of anti-gut bacterial antibody in PBS-T containing 1% BSA and incubated on ice for 1 hour. After washing, FITC-conjugated anti-mouse IgG was added, and the sample was incubated for 30 minutes on ice in the dark. The stained bacteria were then washed and analyzed by flow cytometry¹⁾.

Western Blotting

Phocaeicola vulgatus JCM5826 was collected by centrifugation at 10,000 x g for 5 minutes. Two micrograms of protein were separated by SDS-PAGE, transferred to a PVDF membrane, and blocked. Immunodetection was performed using 25–50 ng/mL of anti-gut bacterial antibody and HRP-conjugated mouse IgG¹⁾.

1. Yoshii, K. et al.: Sci. Rep., 15(1), 1(2025).
   Establishment of enterotype-specific antibodies for various diagnostic systems