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Ff-nano, short functionalized nanorods derived from Ff (f1, fd, or M13) filamentous bacteriophage.

Sattar S, Bennett NJ, Wen WX, Guthrie JM, Blackwell LF, Conway JF, Rakonjac J - Front Microbiol (2015)

Bottom Line: In contrast to standard Ff-derived vectors that replicate in E. coli and contain antibiotic-resistance genes, Ff-nano are protein-DNA complexes that cannot replicate on their own and do not contain any coding sequences.These nanorods show an increased resistance to heating at 70(∘)C in 1% SDS in comparison to the full-length Ff phage of the same coat composition.We demonstrate that functionalized Ff-nano particles are suitable for application as detection particles in sensitive and quantitative "dipstick" lateral flow diagnostic assay for human plasma fibronectin.

View Article: PubMed Central - PubMed

Affiliation: Institute of Fundamental Sciences, Massey University Palmerston North, New Zealand.

ABSTRACT
F-specific filamentous phage of Escherichia coli (Ff: f1, M13, or fd) are long thin filaments (860 nm × 6 nm). They have been a major workhorse in display technologies and bionanotechnology; however, some applications are limited by the high length-to-diameter ratio of Ff. Furthermore, use of functionalized Ff outside of laboratory containment is in part hampered by the fact that they are genetically modified viruses. We have now developed a system for production and purification of very short functionalized Ff-phage-derived nanorods, named Ff-nano, that are only 50 nm in length. In contrast to standard Ff-derived vectors that replicate in E. coli and contain antibiotic-resistance genes, Ff-nano are protein-DNA complexes that cannot replicate on their own and do not contain any coding sequences. These nanorods show an increased resistance to heating at 70(∘)C in 1% SDS in comparison to the full-length Ff phage of the same coat composition. We demonstrate that functionalized Ff-nano particles are suitable for application as detection particles in sensitive and quantitative "dipstick" lateral flow diagnostic assay for human plasma fibronectin.

No MeSH data available.


Related in: MedlinePlus

Fibronectin dipstick assay using Ff-nanoFnB. (A) Schematic representation of a lateral-flow dipstick assay; Fn-detection dipstick assay using: (B) unlabeled; (C) FITC-labeled particles. Each assay (50 μL) contained 1 × 1010 full-length Rnano3 (Rnano3FnB) or 1 × 1011 Ff-nano (or Ff-nanoFnB) particles and 1 μg of Fn. The assay was performed and the unlabeled or FITC-labeled particles were detected as described in Section “Materials and Methods.” The test line, printed with collagen solution, appears as a triple band when the signal is high. This is due to secondary lines flanking the main line that form during printing of collagen on the card. The triple banding during printing is caused by the acidity of the solution, necessary to keep the collagen soluble (0.25% acetic acid).
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Figure 5: Fibronectin dipstick assay using Ff-nanoFnB. (A) Schematic representation of a lateral-flow dipstick assay; Fn-detection dipstick assay using: (B) unlabeled; (C) FITC-labeled particles. Each assay (50 μL) contained 1 × 1010 full-length Rnano3 (Rnano3FnB) or 1 × 1011 Ff-nano (or Ff-nanoFnB) particles and 1 μg of Fn. The assay was performed and the unlabeled or FITC-labeled particles were detected as described in Section “Materials and Methods.” The test line, printed with collagen solution, appears as a triple band when the signal is high. This is due to secondary lines flanking the main line that form during printing of collagen on the card. The triple banding during printing is caused by the acidity of the solution, necessary to keep the collagen soluble (0.25% acetic acid).

Mentions: To investigate the detection of fibronectin (the analyte) in solution using the Ff-nano displaying FnB domains as the detector (probe) in lateral flow devices, a simple dipstick assay was designed and tested (see Figure 5A for a schematic representation of the Fn-detection dipstick assay). The dipsticks used in this assay contained human type I collagen at the test line. Collagen binds fibronectin with high affinity (Engvall et al., 1981).


Ff-nano, short functionalized nanorods derived from Ff (f1, fd, or M13) filamentous bacteriophage.

Sattar S, Bennett NJ, Wen WX, Guthrie JM, Blackwell LF, Conway JF, Rakonjac J - Front Microbiol (2015)

Fibronectin dipstick assay using Ff-nanoFnB. (A) Schematic representation of a lateral-flow dipstick assay; Fn-detection dipstick assay using: (B) unlabeled; (C) FITC-labeled particles. Each assay (50 μL) contained 1 × 1010 full-length Rnano3 (Rnano3FnB) or 1 × 1011 Ff-nano (or Ff-nanoFnB) particles and 1 μg of Fn. The assay was performed and the unlabeled or FITC-labeled particles were detected as described in Section “Materials and Methods.” The test line, printed with collagen solution, appears as a triple band when the signal is high. This is due to secondary lines flanking the main line that form during printing of collagen on the card. The triple banding during printing is caused by the acidity of the solution, necessary to keep the collagen soluble (0.25% acetic acid).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4403547&req=5

Figure 5: Fibronectin dipstick assay using Ff-nanoFnB. (A) Schematic representation of a lateral-flow dipstick assay; Fn-detection dipstick assay using: (B) unlabeled; (C) FITC-labeled particles. Each assay (50 μL) contained 1 × 1010 full-length Rnano3 (Rnano3FnB) or 1 × 1011 Ff-nano (or Ff-nanoFnB) particles and 1 μg of Fn. The assay was performed and the unlabeled or FITC-labeled particles were detected as described in Section “Materials and Methods.” The test line, printed with collagen solution, appears as a triple band when the signal is high. This is due to secondary lines flanking the main line that form during printing of collagen on the card. The triple banding during printing is caused by the acidity of the solution, necessary to keep the collagen soluble (0.25% acetic acid).
Mentions: To investigate the detection of fibronectin (the analyte) in solution using the Ff-nano displaying FnB domains as the detector (probe) in lateral flow devices, a simple dipstick assay was designed and tested (see Figure 5A for a schematic representation of the Fn-detection dipstick assay). The dipsticks used in this assay contained human type I collagen at the test line. Collagen binds fibronectin with high affinity (Engvall et al., 1981).

Bottom Line: In contrast to standard Ff-derived vectors that replicate in E. coli and contain antibiotic-resistance genes, Ff-nano are protein-DNA complexes that cannot replicate on their own and do not contain any coding sequences.These nanorods show an increased resistance to heating at 70(∘)C in 1% SDS in comparison to the full-length Ff phage of the same coat composition.We demonstrate that functionalized Ff-nano particles are suitable for application as detection particles in sensitive and quantitative "dipstick" lateral flow diagnostic assay for human plasma fibronectin.

View Article: PubMed Central - PubMed

Affiliation: Institute of Fundamental Sciences, Massey University Palmerston North, New Zealand.

ABSTRACT
F-specific filamentous phage of Escherichia coli (Ff: f1, M13, or fd) are long thin filaments (860 nm × 6 nm). They have been a major workhorse in display technologies and bionanotechnology; however, some applications are limited by the high length-to-diameter ratio of Ff. Furthermore, use of functionalized Ff outside of laboratory containment is in part hampered by the fact that they are genetically modified viruses. We have now developed a system for production and purification of very short functionalized Ff-phage-derived nanorods, named Ff-nano, that are only 50 nm in length. In contrast to standard Ff-derived vectors that replicate in E. coli and contain antibiotic-resistance genes, Ff-nano are protein-DNA complexes that cannot replicate on their own and do not contain any coding sequences. These nanorods show an increased resistance to heating at 70(∘)C in 1% SDS in comparison to the full-length Ff phage of the same coat composition. We demonstrate that functionalized Ff-nano particles are suitable for application as detection particles in sensitive and quantitative "dipstick" lateral flow diagnostic assay for human plasma fibronectin.

No MeSH data available.


Related in: MedlinePlus