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A low-cost, high-performance system for fluorescence lateral flow assays.

Lee LG, Nordman ES, Johnson MD, Oldham MF - Biosensors (Basel) (2013)

Bottom Line: A wide range of values for the ratio of signal to nonspecific binding was found, from 50 for R-phycoerythrin (R-PE) to 0.15 for Brilliant Violet 605.Fluorescence detection with R-PE and absorbance detection with colloidal gold were directly compared in lateral flow using biotinylated bovine serum albumen (BSA) as the analyte.We believe our inexpensive yet high-performance platform will be useful for providing quantitative and sensitive detection in a point-of-care setting.

View Article: PubMed Central - PubMed

Affiliation: Song Diagnostic Research LLC, 1 Megans Lane, Woodside, CA 94062, USA. Linda@songdx.net.

ABSTRACT
We demonstrate a fluorescence lateral flow system that has excellent sensitivity and wide dynamic range. The illumination system utilizes an LED, plastic lenses and plastic and colored glass filters for the excitation and emission light. Images are collected on an iPhone 4. Several fluorescent dyes with long Stokes shifts were evaluated for their signal and nonspecific binding in lateral flow. A wide range of values for the ratio of signal to nonspecific binding was found, from 50 for R-phycoerythrin (R-PE) to 0.15 for Brilliant Violet 605. The long Stokes shift of R-PE allowed the use of inexpensive plastic filters rather than costly interference filters to block the LED light. Fluorescence detection with R-PE and absorbance detection with colloidal gold were directly compared in lateral flow using biotinylated bovine serum albumen (BSA) as the analyte. Fluorescence provided linear data over a range of 0.4-4,000 ng/mL with a 1,000-fold signal change while colloidal gold provided non-linear data over a range of 16-4,000 ng/mL with a 10-fold signal change. A comparison using human chorionic gonadotropin (hCG) as the analyte showed a similar advantage in the fluorescent system. We believe our inexpensive yet high-performance platform will be useful for providing quantitative and sensitive detection in a point-of-care setting.

No MeSH data available.


Related in: MedlinePlus

Alexa Fluor 532 has a good ratio of signal to nonspecific binding (S/NSB) compared to Atto 430LS. Each fluorophore is conjugated to streptavidin, spotted on nitrocellulose and the signal read in the breadboard (signal, blue diamonds). Strips of nitrocellulose that have been blocked with BSA are immersed in each solution and read in the breadboard (nonspecific binding, magenta squares). The ratios of the two slopes are reported as the S/NSB ratio.
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biosensors-03-00360-f004: Alexa Fluor 532 has a good ratio of signal to nonspecific binding (S/NSB) compared to Atto 430LS. Each fluorophore is conjugated to streptavidin, spotted on nitrocellulose and the signal read in the breadboard (signal, blue diamonds). Strips of nitrocellulose that have been blocked with BSA are immersed in each solution and read in the breadboard (nonspecific binding, magenta squares). The ratios of the two slopes are reported as the S/NSB ratio.

Mentions: Shown in Figure 4 are the signal and NSB data for two of the dyes, Alexa Fluor 532 and Atto 430LS. Table 2 shows the ratio of signal to NSB for all the dyes that we analyzed. Surprisingly, even though all the dyes were very water-soluble, they showed a wide range in ratios of signal to NSB. Brilliant Violet 605 streptavidin was extraordinarily “sticky”, actually producing greater signal in the nonspecific binding mode than the signal mode for each dilution of dye-labeled streptavidin. Alexa Fluor 532 streptavidin and R-PE streptavidin were the least sticky. It is clear from these results that in addition to the inherent brightness of a fluorescent dye, the ratio of signal to NSB is a key characteristic in determining the utility of a dye in lateral flow. A dye with a high ratio of signal to NSB will have a good dynamic range, since high concentrations of dye can be used to saturate high concentrations of analyte without causing too much background for low concentrations of analyte.


A low-cost, high-performance system for fluorescence lateral flow assays.

Lee LG, Nordman ES, Johnson MD, Oldham MF - Biosensors (Basel) (2013)

Alexa Fluor 532 has a good ratio of signal to nonspecific binding (S/NSB) compared to Atto 430LS. Each fluorophore is conjugated to streptavidin, spotted on nitrocellulose and the signal read in the breadboard (signal, blue diamonds). Strips of nitrocellulose that have been blocked with BSA are immersed in each solution and read in the breadboard (nonspecific binding, magenta squares). The ratios of the two slopes are reported as the S/NSB ratio.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263565&req=5

biosensors-03-00360-f004: Alexa Fluor 532 has a good ratio of signal to nonspecific binding (S/NSB) compared to Atto 430LS. Each fluorophore is conjugated to streptavidin, spotted on nitrocellulose and the signal read in the breadboard (signal, blue diamonds). Strips of nitrocellulose that have been blocked with BSA are immersed in each solution and read in the breadboard (nonspecific binding, magenta squares). The ratios of the two slopes are reported as the S/NSB ratio.
Mentions: Shown in Figure 4 are the signal and NSB data for two of the dyes, Alexa Fluor 532 and Atto 430LS. Table 2 shows the ratio of signal to NSB for all the dyes that we analyzed. Surprisingly, even though all the dyes were very water-soluble, they showed a wide range in ratios of signal to NSB. Brilliant Violet 605 streptavidin was extraordinarily “sticky”, actually producing greater signal in the nonspecific binding mode than the signal mode for each dilution of dye-labeled streptavidin. Alexa Fluor 532 streptavidin and R-PE streptavidin were the least sticky. It is clear from these results that in addition to the inherent brightness of a fluorescent dye, the ratio of signal to NSB is a key characteristic in determining the utility of a dye in lateral flow. A dye with a high ratio of signal to NSB will have a good dynamic range, since high concentrations of dye can be used to saturate high concentrations of analyte without causing too much background for low concentrations of analyte.

Bottom Line: A wide range of values for the ratio of signal to nonspecific binding was found, from 50 for R-phycoerythrin (R-PE) to 0.15 for Brilliant Violet 605.Fluorescence detection with R-PE and absorbance detection with colloidal gold were directly compared in lateral flow using biotinylated bovine serum albumen (BSA) as the analyte.We believe our inexpensive yet high-performance platform will be useful for providing quantitative and sensitive detection in a point-of-care setting.

View Article: PubMed Central - PubMed

Affiliation: Song Diagnostic Research LLC, 1 Megans Lane, Woodside, CA 94062, USA. Linda@songdx.net.

ABSTRACT
We demonstrate a fluorescence lateral flow system that has excellent sensitivity and wide dynamic range. The illumination system utilizes an LED, plastic lenses and plastic and colored glass filters for the excitation and emission light. Images are collected on an iPhone 4. Several fluorescent dyes with long Stokes shifts were evaluated for their signal and nonspecific binding in lateral flow. A wide range of values for the ratio of signal to nonspecific binding was found, from 50 for R-phycoerythrin (R-PE) to 0.15 for Brilliant Violet 605. The long Stokes shift of R-PE allowed the use of inexpensive plastic filters rather than costly interference filters to block the LED light. Fluorescence detection with R-PE and absorbance detection with colloidal gold were directly compared in lateral flow using biotinylated bovine serum albumen (BSA) as the analyte. Fluorescence provided linear data over a range of 0.4-4,000 ng/mL with a 1,000-fold signal change while colloidal gold provided non-linear data over a range of 16-4,000 ng/mL with a 10-fold signal change. A comparison using human chorionic gonadotropin (hCG) as the analyte showed a similar advantage in the fluorescent system. We believe our inexpensive yet high-performance platform will be useful for providing quantitative and sensitive detection in a point-of-care setting.

No MeSH data available.


Related in: MedlinePlus