Optical microchips based on high-affinity recombinant protein binders-Human serum albumin detection in urine

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F44555601%3A13440%2F18%3A43894070" target="_blank" >RIV/44555601:13440/18:43894070 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/86652036:_____/18:00495868 RIV/61388971:_____/18:00495868

Výsledek na webu

<a href="https://www.sciencedirect.com/science/article/pii/S0925400518310980" target="_blank" >https://www.sciencedirect.com/science/article/pii/S0925400518310980</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1016/j.snb.2018.05.180" target="_blank" >10.1016/j.snb.2018.05.180</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Optical microchips based on high-affinity recombinant protein binders-Human serum albumin detection in urine

Popis výsledku v původním jazyce

Recent developments in molecular evolution technologies have led to novel types of high-affinity recombinant protein binders (PB) able to substitute antibodies in many diagnostic and therapeutic applications. Despite almost a decade of research, they have so far only been sporadically used for biosensor construction. Here, we present a proof-of-principle comparative study focused on the application of three types of PB recognizing human serum albumin (HSA) in the fabrication of diagnostic optical microchips detecting clinically relevant HSA levels in urine. The PB tested were: (i) biotinylated anti-HSA Affibody (AF) (IgG binding domain of protein A, Staphylococcus aureus); (ii) biotinylated protein construct based on albumin-binding domain (ABD) of protein G (Streptococcus G148) fused with long TolA spacer (6xHis-WT-ABD-TolA-AviTag) and (iii) WT-ABD-Trp leaderstreptavidin tetrameric fusion protein (SA-ABD-WT). Open glass microchips with 24 independent microwells (volume 8 ?L) and micropatterned detection zones were prepared and used for oriented binding of proteins through the biotin/streptavidin chemistry. The analytical performance of the optical microchips was tested by performing direct specific detection of fluorescently labelled HSA in various environments. Results show that the length of peptide spacer present between the binding protein domain and sensor surface is a key factor influencing biosensor performance. The biosensor based on SA-ABD-WT reached the limit of detection (LOD) for HSA in urine (LOD=0.65 ?g/ml) sufficient to identify the chronic kidney disease caused by high blood pressure or diabetes. Furthermore, it offers the highest signal intensity, low noise and significant simplification of microchip preparation due to a simple one-step immobilization procedure. Our results may be further exploited in development of diagnostic microchips dedicated to the detection of a wide range of molecular targets recognized by specific ABD protein binders.

Název v anglickém jazyce

Optical microchips based on high-affinity recombinant protein binders-Human serum albumin detection in urine

Popis výsledku anglicky

Recent developments in molecular evolution technologies have led to novel types of high-affinity recombinant protein binders (PB) able to substitute antibodies in many diagnostic and therapeutic applications. Despite almost a decade of research, they have so far only been sporadically used for biosensor construction. Here, we present a proof-of-principle comparative study focused on the application of three types of PB recognizing human serum albumin (HSA) in the fabrication of diagnostic optical microchips detecting clinically relevant HSA levels in urine. The PB tested were: (i) biotinylated anti-HSA Affibody (AF) (IgG binding domain of protein A, Staphylococcus aureus); (ii) biotinylated protein construct based on albumin-binding domain (ABD) of protein G (Streptococcus G148) fused with long TolA spacer (6xHis-WT-ABD-TolA-AviTag) and (iii) WT-ABD-Trp leaderstreptavidin tetrameric fusion protein (SA-ABD-WT). Open glass microchips with 24 independent microwells (volume 8 ?L) and micropatterned detection zones were prepared and used for oriented binding of proteins through the biotin/streptavidin chemistry. The analytical performance of the optical microchips was tested by performing direct specific detection of fluorescently labelled HSA in various environments. Results show that the length of peptide spacer present between the binding protein domain and sensor surface is a key factor influencing biosensor performance. The biosensor based on SA-ABD-WT reached the limit of detection (LOD) for HSA in urine (LOD=0.65 ?g/ml) sufficient to identify the chronic kidney disease caused by high blood pressure or diabetes. Furthermore, it offers the highest signal intensity, low noise and significant simplification of microchip preparation due to a simple one-step immobilization procedure. Our results may be further exploited in development of diagnostic microchips dedicated to the detection of a wide range of molecular targets recognized by specific ABD protein binders.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

20602 - Medical laboratory technology (including laboratory samples analysis; diagnostic technologies) (Biomaterials to be 2.9 [physical characteristics of living material as related to medical implants, devices, sensors])

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)<br>S - Specificky vyzkum na vysokych skolach

Rok uplatnění

2018

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Sensors and Actuators. B - Chemical

ISSN

0925-4005

e-ISSN

—

Svazek periodika

2018

Číslo periodika v rámci svazku

272

Stát vydavatele periodika

CH - Švýcarská konfederace

Počet stran výsledku

7

Strana od-do

441-447

Kód UT WoS článku

000439715000051

EID výsledku v databázi Scopus

2-s2.0-85048547014