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Abstract
A rapid and sensitive detection technology is highly desirable for specific detection of E. coli O157:H7, one of the leading bacterial pathogens causing foodborne illness. In this study, we reported the rapid detection of E. coli O157:H7 by using calcium signaling of the B cell upon cellular membrane anchors anti-E. coli O157:H7 IgM. The binding of E. coli O157:H7 to the IgM on B cell surface activates the B cell receptor (BCR)-induced Ca2+ signaling pathway and results in the release of Ca2+ within seconds. The elevated intracellular Ca2+ triggers Fura-2, a fluorescent Ca2+ indicator, for reporting the presence of pathogens. The Fura-2 is transferred to B cells before detection. The study demonstrated that the developed B cell based biosensor was able to specifically detect E. coli O157:H7 at the low concentration within 10 min in pure culture samples. Finally, the B cell based biosensor was used for the detection of E. coli O157:H7 in ground beef samples. With its short detection time and high sensitivity at the low concentration of the target bacteria, this B cell biosensor shows promise in future application of the high throughput and rapid food detection, biosafety and environmental monitoring.
Highlights
There are estimated 48 million cases of foodborne illness resulting in 3,000 deaths, and an estimated cost of 78 billion dollars per year[1,2,3]
Combining advantages of the sensitivity of Fura-2 to Ca2+ and the rapidness of response of B cell to antigens, a Ca2+-indicator based B cell biosensor was designed for E. coli O157:H7 detection
The B cell receptor (BCR) is a multisubunit protein complex composed of a membrane form of immunoglobulin (Ig) that is noncovalently associated with heterodimers of Igα and Igβ
Summary
There are estimated 48 million cases of foodborne illness resulting in 3,000 deaths, and an estimated cost of 78 billion dollars per year[1,2,3]. A series of fluorescent calcium indicator dyes have been developed for measurement of free intracellular calcium in eukaryotic cells and prokaryote. Fura-2 has been known as an indicator dye for measuring the concentration of free calcium ([Ca2+]i) within living cells[30,31]. The ratio of fluorescence emission at the two excitation wave lengths (340:380) is considered a reliable indicator of [Ca2+]i32,33. It has been widely used but not limited in immunology, cytology and neurology for interrogating ion channels. The innovative approach in this study is the use of a Ca2+-indicator, Fura-2 for detecting BCR-induced Ca2+ change due to an interaction between B cells and pathogens. It was found that a fluorescence ratio at 340/380 is correlated to a concentration of free intracellular Ca2+30.This technology has a great potential to provide a practical alternative for detection of E. coli O157:H7 and other pathogens
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