Streptavidin Coverage on Biotinylated Surfaces

Abstract

Biosensors and other biological platform technologies require the functionalization of their surface with receptors to enhance affinity and selectivity. Control over the functionalization density is required to tune the platform’s properties. Streptavidin (SAv) monolayers are widely used to immobilize biotinylated proteins, receptors, and DNA. The SAv density on a surface can be varied easily, but the predictability is dependent on the method by which the SAv is immobilized. In this study we show a method to quantitatively predict the SAv coverage on biotinylated surfaces. The method is validated by measuring the SAv coverage on supported lipid bilayers with a range of biotin contents and two different main phase lipids and by using quartz crystal microbalance and localized surface plasmon resonance. We explore a predictive model of the biotin-dependent SAv coverage without any fit parameters. Model and data allow to predict the SAv coverage based on the biotin coverage, in both the low- and high-density regimes. This is of special importance in applications with multivalent binding where control over surface receptor density is required, but a direct measurement is not possible.

Distinct Binding Properties of Neutravidin and Streptavidin Proteins to Biotinylated Supported Lipid Bilayers: Implications for Sensor Functionalization

“…Regarding the applicability of these scientific results to various SLB platforms, it should be noted that the results were obtained using a zwitterionic SLB, which is widely known to have antifouling properties [ 19 ] and is hence routinely used. Depending on the sensing application or other biophysical objective, it is possible to change the lipid composition, which might affect the protein attachment specifics.…”

“…One emerging use of avidin–biotin technology is in the field of supported lipid bilayer (SLB) coatings [ 12 ], which are antifouling cell membrane mimics that are formed through the self-assembly of phospholipid molecules on solid surfaces and are widely used in biosensor development [ 13 , 14 , 15 , 16 ]. Generally, biotinylated lipids are incorporated in the SLB that acts as a platform upon which avidin or related protein analogues are specifically attached via biotin coupling, and such platforms can be used in various biosensing applications ranging from fundamental biophysical studies to molecular diagnostics and cancer cell detection [ 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 , 26 , 27 , 28 ] ( Figure 1 A).…”

“…Sensors 2022, 22, x FOR PEER REVIEW 2 of 15 solid surfaces and are widely used in biosensor development [13][14][15][16]. Generally, biotinylated lipids are incorporated in the SLB that acts as a platform upon which avidin or related protein analogues are specifically attached via biotin coupling, and such platforms can be used in various biosensing applications ranging from fundamental biophysical studies to molecular diagnostics and cancer cell detection [17][18][19][20][21][22][23][24][25][26][27][28] (Figure 1A). Avidin is an egg-white glycoprotein that has four binding sites for biotin along with a high isoelectric point (pI) around pH 10.5 (ref.…”

Unraveling pH Effects on Ultrafiltration Membrane Fouling by Extracellular Polymeric Substances: Adsorption and Conformation Analyzed with Localized Surface Plasmon Resonance

“…The adherence and adsorption kinetics of the extracted EPS were determined by using the modified QCM-D−LSPR sensors of Insplorion AB (Figure 1) mounted in QCM-D flow cell with a sapphire window hosted inside the Insplorion Acoulyte module. 45 Prior to each measurement, the crystals were sonicated for 10 min in 2% sodium dodecyl sulfate (SDS) followed by a rinse with double distilled water. Then, the sensors were sonicated for 10 min in 100% isopropanol, and finally the sensors were rinsed thoroughly with double distilled water and dried with 99.99% N 2 .…”

Density Control over MBD2 Receptor-Coated Surfaces Provides Superselective Binding of Hypermethylated DNA

“…60 In comparison, a densely packed streptavidin layer typically shows a frequency shift of around 25 Hz. 61 Even though streptavidin and MBD2 have different molecular weights and their degrees of hydration (which affects QCM frequencies) may differ as well, we regard the here observed MBD2 frequency values ranging from 35 Hz (279 ng/cm 2 ) to 55 Hz (438 ng/cm 2 ) as indicative of dense packing because the frequency values level off in this range.…”