Bio-layer interferometry: Difference between revisions

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{{Short description|Optical biosensing technology}}

[[File:Bio-layer interferometry without analyte binding.gif|thumb|Figure 1 - Overview schematic of a Bio-layer interferometry setup|300x300px]][[File:Thin film interference - soap bubble.gif|thumb|265x265px|Figure 2 - The ligand-analyte layer creates an optical path length difference, reflecting incident light in two different patterns]]'''Bio-layer interferometry''' ('''BLI)''') is an optical biosensing technology that analyzes biomolecular interactions in real-time without the need for fluorescent labeling.<ref name="Apiyo_2017">{{Cite book| vauthors = Apiyo D, Schasfoort R, Schuck P, Marquart A, Gedig ET, Karlsson R, Abdiche YN, Eckman Y, Blum SR, Schasfoort RB |title=Handbook of Surface Plasmon Resonance.|date=2017|publisher=Royal Society of Chemistry|isbn=978-1-78801-139-6|oclc=988866146}}</ref> Alongside [[Surfacesurface plasmon resonance|Surface Plasmon Resonance]] (SPR), BLI is one of few widely available [[Label-free quantification|label-free]] biosensing technologies, a detection style that yields more information in less time than traditional processes.<ref>{{cite journal | vauthors = Syahir A, Usui K, Tomizaki KY, Kajikawa K, Mihara H | title = Label and Label-Free Detection Techniques for Protein Microarrays | journal = Microarrays | volume = 4 | issue = 2 | pages = 228–244 | date = April 2015 | pmid = 27600222 | pmc = 4996399 | doi = 10.3390/microarrays4020228 | doi-access = free }}</ref> The technology relies on the phase shift-wavelength correlation created between interference patterns off of two unique surfaces on the tip of a biosensor.<ref name=":1">{{cite journal | vauthors = Müller-Esparza H, Osorio-Valeriano M, Steube N, Thanbichler M, Randau L | title = Bio-Layer Interferometry Analysis of the Target Binding Activity of CRISPR-Cas Effector Complexes | journal = Frontiers in Molecular Biosciences | volume = 7 | pages = 98 | date = 2020-05-27 | pmid = 32528975 | pmc = 7266957 | doi = 10.3389/fmolb.2020.00098 | doi-access = free }}</ref> BLI has significant applications in quantifying binding strength, measuring protein interactions, and identifying properties of reaction kinetics, such as rate constants and reaction rates.<ref>{{cite journal | vauthors = Rich RL, Myszka DG | title = Higher-throughput, label-free, real-time molecular interaction analysis | journal = Analytical Biochemistry | volume = 361 | issue = 1 | pages = 1–6 | date = February 2007 | pmid = 17145039 | doi = 10.1016/j.ab.2006.10.040 }}</ref>

== Method ==

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=== Mechanism overview ===

[[File:Bio-layer interferometry wavelength shift due to analyte binding.gif|thumb|Figure 3 - Reflectance signal as a function of wavelength]]

Bio-layer interferometry measures kinetics and biomolecular interactions on a basis of [[wave interference]]. To prepare for BLI analysis between two unique biomolecules, the ligand is first immobilized onto a bio compatible [[biosensor]] while the [[analyte]] is in solution.<ref name=":22">{{cite journal | vauthors = Müller-Esparza H, Osorio-Valeriano M, Steube N, Thanbichler M, Randau L | title = Bio-Layer Interferometry Analysis of the Target Binding Activity of CRISPR-Cas Effector Complexes | journal = Frontiers in Molecular Biosciences | volume = 7 | pages = 98 | date = 2020-05-27 | pmid = 32528975 | doi = 10.3389/fmolb.2020.00098 | pmc = 7266957 | doi-access = free }}</ref> Shortly after this, the biosensor tip is dipped into the solution and the target molecule will begin to associate with the analyte, producing a layer on top of the biosensor tip. This creates two separate surfaces: i) the molecule immobilized on the biosensor tipsubstrate itself, and ii) the substrate thatinteracting iswith boundthe tomolecule itimmobilized fromon the solutionbiosensor tip.<ref name="Apiyo_2017" /> This essentially creates a [[thin-film interference]], in which the created layer acts as a thin film bound by these two surfaces. White light from a tungsten lamp is shone onto the biosensor tip and reflected off both surfaces, creating two unique reflection patterns with different [[Luminous intensity|intensities]].<ref name=":22" /> Figure 2 expresses this phenomenon in a more general form. The wavelength shift (Δλ) between these two reflection patterns creates an interference pattern (Figure 3) from which all desired results can be obtained.<ref name="Apiyo_2017" /> Since the wavelength shift is direct measure of the change in thickness of the biological layer and the biological layer thickness will change in response to molecules associating to and dissociating from the biosensor, the interference pattern will allow for real-time monitoring of molecular interactions on the biosensor surface.<ref name=":13">{{cite journal | vauthors = Wallner J, Lhota G, Jeschek D, Mader A, Vorauer-Uhl K | title = Application of Bio-Layer Interferometry for the analysis of protein/liposome interactions | journal = Journal of Pharmaceutical and Biomedical Analysis | volume = 72 | pages = 150–154 | date = January 2013 | pmid = 23146240 | doi = 10.1016/j.jpba.2012.10.008 }}</ref> In short, a positive wavelength shift implies an increase in biolayer thickness and thus more association, while a negative wavelength shift implies a decrease in biolayer thickness and thus more dissociation.<ref name=":13" />

=== "Dip and read" format ===

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=== Biosensor type and selection ===

Bio-layer interferometry relies on [[Biosensor|biosensors]] with a fiber optic tip upon which the ligand is immobilized.<ref name="Apiyo_2017" /> The tip is additionally coated with a matrix biocompatible with the target molecule to limit any non-specific binding. For BLI calculations to work, it is necessary to assume that both the fiber optic tip and the bound ligand and analyte act as thin, reflective surfaces.<ref>{{Cite journal| vauthors = Gao S, Zheng X, Wu J |date=2017|title=A biolayer interferometry-based competitive biosensor for rapid and sensitive detection of saxitoxin |journal=Sensors and Actuators B: Chemical|volume=246|pages=169–174|doi=10.1016/j.snb.2017.02.078|bibcode=2017SeAcB.246..169G |issn=0925-4005}}</ref> The biosensors are disposable, resulting in low costs and high commercial availability.<ref>{{cite journal | vauthors = Abdiche Y, Malashock D, Pinkerton A, Pons J | title = Determining kinetics and affinities of protein interactions using a parallel real-time label-free biosensor, the Octet | journal = Analytical Biochemistry | volume = 377 | issue = 2 | pages = 209–217 | date = June 2008 | pmid = 18405656 | doi = 10.1016/j.ab.2008.03.035 | doi-access = free }}</ref> Biosensor selection is determined by the desired test results: kinetic analysis, quantitative analysis, or both.<ref>{{cite journal | vauthors = Yu Y, Mitchell S, Lynaugh H, Brown M, Nobrega RP, Zhi X, Sun T, Caffry I, Cao Y, Yang R, Burnina I, Xu Y, Estep P | display-authors = 6 | title = Understanding ForteBio's Sensors for High-Throughput Kinetic and Epitope Screening for Purified Antibodies and Yeast Culture Supernatant | journal = Journal of Biomolecular Screening | volume = 21 | issue = 1 | pages = 88–95 | date = January 2016 | pmid = 26442912 | doi = 10.1177/1087057115609564 | pmc = 4708621 | doi-access = free }}</ref> Most commercially available biosensor types will be grouped into one of these three categories by the BLI manufacturer.<ref name="Apiyo_2017" />

== Applications ==