Gold Nanoparticle Paper Immunoassays for Sensing the Presence of Vibrio parahaemolyticus in Oyster Hemolymph

Abstract

Seafood contamination with Vibrio bacteria is a problem for aquaculture, especially with oysters, which are often consumed raw. Current methods for diagnosing bacterial pathogens in seafood involve lab-based assays such as polymerase chain reaction or culturing, which are time consuming and must occur in a centralized location. Detection of Vibrio in a point-of-care assay would be a significant tool for food safety control measures. We report here a paper immunoassay that can detect the presence of Vibrio parahaemolyticus (Vp) in buffer and oyster hemolymph. The test uses gold nanoparticles conjugated to polyclonal anti-Vibrio antibodies in a paper-based sandwich immunoassay. A sample is added to the strip and wicked through by capillary action. If Vp is present, it results in a visible color at the test area that can be read out by eyes or a standard mobile phone camera. The assay has a limit of detection of 6.05 × 10 5 cfu/mL and a cost estimate of $5 per test. Receiver operating characteristic curves with validated environmental samples showed a test sensitivity of 0.96 and a specificity of 1.00. Because the assay is inexpensive and can be used on Vp directly without the requirement for culturing, or sophisticated equipment, it has the potential to be used in fieldable settings.

Paper-Powered Synthesis of Gold Nanoparticles and Metronidazole for Bioassays

“…17 Thus, functionality in an immunoassay is a good test for the NP biocompatibility, bioconjugation, and stability. 24,25 Extracted Ampli-synthesized NPs were conjugated to anti-IgG antibodies by physisorption 16 followed by a PEG backfill (Supporting Information Figure S6). 26 Immunoassays consisted of a nitrocellulose strip with anti-IgG spotted at the test area and anti-Fc spotted at the control area to bind to the NPanti-IgG directly.…”

“…Immunoassays are highly sensitive to NP aggregation due to interface effects, where nonspecific adsorption can prevent proper binding of the target to the NP-Ab or immobilized antibody, resulting in smearing, high background intensities, false positives, or no signal due to precipitation in the dipstick solution . Thus, functionality in an immunoassay is a good test for the NP biocompatibility, bioconjugation, and stability. , …”