Detection of cholera toxin with surface plasmon field-enhanced fluorescent spectroscopy


Seherler S., Bozdogan A., Özal Ildeniz T. A. , Kök F. N. , Sakir I. A.

BIOTECHNOLOGY AND APPLIED BIOCHEMISTRY, 2021 (Journal Indexed in SCI) identifier identifier identifier

  • Publication Type: Article / Article
  • Volume:
  • Publication Date: 2021
  • Doi Number: 10.1002/bab.2227
  • Title of Journal : BIOTECHNOLOGY AND APPLIED BIOCHEMISTRY
  • Keywords: antibody, biosensor, cholera toxin, environmental biotechnology, immobilization, surface plasmon resonance (SPR), surface plasmon field-enhanced fluorescent spectroscopy (SPFS), QUARTZ-CRYSTAL MICROBALANCE, VIBRIO-CHOLERAE, SENSITIVE DETECTION, RAPID DETECTION, FOOD SAFETY, RESONANCE, BIOSENSOR, WATER, IMMUNOSENSOR, PATHOGENS

Abstract

In this work, a biosensor based on surface plasmon field-enhanced florescence spectroscopy (SPFS) method was successfully constructed to detect the truncated form of cholera toxin, that is, its beta subunit (CTX-B). CTX-B is a relatively small molecule (12 kDa) and it was chosen as model analyte for the detection of protein toxins originated from waterborne pathogens. Recognition layer was prepared on gold-coated LaSFN9 glasses modified with 11-mercaptoundecanoic acid (11-MUA). Biotin-conjugated anti-CTX-B polyclonal antibody (B-Ab) was immobilized on streptavidin (SA) layer constructed on the 11-MUA-modified surface. CTX-B amount was determined with direct assay using B-Ab in surface plasmon resonance (SPR) mode and with sandwich assay in SPFS mode using Cy5-conjugated anti-CTX-B polyclonal antibody. Minimum detected CTX-B concentrations were 10 and 0.01 mu g/ml with SPR and SPFS, respectively, showing the sensitivity of the SPFS system over the conventional one. The detection was done in 2-6 h, which was faster than both culture and polymerase chain reaction (PCR)-based methods. Stability tests were performed with SA-coated sensors (excluding B-Ab). In this form, the layer was stable after 30 days of storage in phosphate-buffered saline (PBS; 0.01 M, pH = 7.4) at +4 degrees C. B-Ab layer was formed immediately on them before each measurement.