Fluorescence immunoassay of E. coli using anti-lipopolysaccharide antibodies isolated from human serum

Ji Hong Bong, Jiyun Kim, Ga Yeon Lee, Jun Hee Park, Tae Hun Kim, Min Jung Kang, Jae Chul Pyun

Research output: Contribution to journalArticle

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Abstract

In this work, the gram-negative bacterium Escherichia coli strain BL21(DE3) (with lipopolysaccharide (LPS) in its outer membrane) and its modified ClearColi™ strain (lacking LPS) were used for the separation of anti-LPS antibodies from human serum by the following steps: (1) binding of the serum proteins to BL21(DE3); (2) dissociation of the bound proteins (including anti-LPS antibodies) from BL21(DE3) with acid; (3) filtering of the dissociated proteins using ClearColi to remove unwanted proteins; and (4) separation of the antibody fraction by protein-A column chromatography. The binding properties of the separated antibodies were analyzed by fluorescence-activated cell sorting to confirm their selective binding to LPS on the outer membrane of BL21(DE3), and by thermophoretic immunoassay to estimate their dissociation constant. The in vitro applicability of the separated anti-LPS antibodies was demonstrated through a fluorescence assay of BL21(DE3), after immobilizing the antibodies onto a modified microplate surface. The electrochemical detection of BL21(DE3) was also achieved after immobilizing the anti-LPS antibodies onto a gold electrode.

Original languageEnglish
Pages (from-to)518-528
Number of pages11
JournalBiosensors and Bioelectronics
Volume126
DOIs
Publication statusPublished - 2019 Feb 1

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Immunoassay
Antibodies
Escherichia coli
Lipopolysaccharides
Anti-Idiotypic Antibodies
Fluorescence
Serum
Proteins
Membranes
Column chromatography
Staphylococcal Protein A
Gram-Negative Bacteria
Sorting
Gold
Chromatography
Blood Proteins
Assays
Bacteria
Carrier Proteins
Flow Cytometry

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Biophysics
  • Biomedical Engineering
  • Electrochemistry

Cite this

Bong, Ji Hong ; Kim, Jiyun ; Lee, Ga Yeon ; Park, Jun Hee ; Kim, Tae Hun ; Kang, Min Jung ; Pyun, Jae Chul. / Fluorescence immunoassay of E. coli using anti-lipopolysaccharide antibodies isolated from human serum. In: Biosensors and Bioelectronics. 2019 ; Vol. 126. pp. 518-528.
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Fluorescence immunoassay of E. coli using anti-lipopolysaccharide antibodies isolated from human serum. / Bong, Ji Hong; Kim, Jiyun; Lee, Ga Yeon; Park, Jun Hee; Kim, Tae Hun; Kang, Min Jung; Pyun, Jae Chul.

In: Biosensors and Bioelectronics, Vol. 126, 01.02.2019, p. 518-528.

Research output: Contribution to journalArticle

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