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Label-free biomarker detection from whole blood.

Stern E, Vacic A, Rajan NK, Criscione JM, Park J, Ilic BR, Mooney DJ, Reed MA, Fahmy TM - Nat Nanotechnol (2009)

Bottom Line: However, detecting these biomarkers in physiological fluid samples is difficult because of problems such as biofouling and non-specific binding, and the resulting need to use purified buffers greatly reduces the clinical relevance of these sensors.This two-stage approach isolates the detector from the complex environment of whole blood, and reduces its minimum required sensitivity by effectively pre-concentrating the biomarkers.This study marks the first use of label-free nanosensors with physiological solutions, positioning this technology for rapid translation to clinical settings.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, School of Engineering and Applied Science, Yale University, New Haven, Connecticut 06511, USA.

ABSTRACT
Label-free nanosensors can detect disease markers to provide point-of-care diagnosis that is low-cost, rapid, specific and sensitive. However, detecting these biomarkers in physiological fluid samples is difficult because of problems such as biofouling and non-specific binding, and the resulting need to use purified buffers greatly reduces the clinical relevance of these sensors. Here, we overcome this limitation by using distinct components within the sensor to perform purification and detection. A microfluidic purification chip simultaneously captures multiple biomarkers from blood samples and releases them, after washing, into purified buffer for sensing by a silicon nanoribbon detector. This two-stage approach isolates the detector from the complex environment of whole blood, and reduces its minimum required sensitivity by effectively pre-concentrating the biomarkers. We show specific and quantitative detection of two model cancer antigens from a 10 microl sample of whole blood in less than 20 min. This study marks the first use of label-free nanosensors with physiological solutions, positioning this technology for rapid translation to clinical settings.

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Related in: MedlinePlus

Schematic of MPC operationa, Primary antibodies to multiple biomarkers, here prostate specific antigen (PSA) and carbohydrate antigen-15.3 (CA15.3), are bound with a photocleavable crosslinker to the MPC. The chip is placed in a plastic housing and a valve (pink) directs fluid flow exiting the chip to either a waste receptacle or the nanosensor chip. b, Whole blood is injected into the chip with the valve set to the waste compartment (black arrow shows the direction of fluid flow) and, if present in the sample, biomarkers bind their cognate antibodies. c, Washing steps follow blood flow and the chip volume (5 μL) is filled with sensing buffer prior to UV irradiation (orange arrows). During UV exposure, the photolabile crosslinker cleaves, releasing the antibody-antigen complexes into solution. d, The valve is set to the nanosensor reservoir (black arrow shows the direction of fluid flow) and the 5 μL volume is transferred, enabling label-free sensing to be performed to determine the presence of specific biomarkers.
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Figure 1: Schematic of MPC operationa, Primary antibodies to multiple biomarkers, here prostate specific antigen (PSA) and carbohydrate antigen-15.3 (CA15.3), are bound with a photocleavable crosslinker to the MPC. The chip is placed in a plastic housing and a valve (pink) directs fluid flow exiting the chip to either a waste receptacle or the nanosensor chip. b, Whole blood is injected into the chip with the valve set to the waste compartment (black arrow shows the direction of fluid flow) and, if present in the sample, biomarkers bind their cognate antibodies. c, Washing steps follow blood flow and the chip volume (5 μL) is filled with sensing buffer prior to UV irradiation (orange arrows). During UV exposure, the photolabile crosslinker cleaves, releasing the antibody-antigen complexes into solution. d, The valve is set to the nanosensor reservoir (black arrow shows the direction of fluid flow) and the 5 μL volume is transferred, enabling label-free sensing to be performed to determine the presence of specific biomarkers.

Mentions: Figure 1 schematically illustrates the operation of the MPC chip. The avidin-functionalized chip19 (Fig. 1a) is treated with antibodies to any number of specific biomarkers conjugated to biotinylated, photocleavable crosslinkers containing a specific 19-mer DNA sequence (Fig. 2a)20. The MPC geometry was chosen to optimize biomarker binding (Supplementary Fig. S1)14 and chips were fabricated from 4-inch silicon wafers in a one-step photolithographic process (Supplementary Fig. S2). Completed chips (Fig. 2b) were loaded into a custom-machined flow chamber (inset, Fig. 2b and Supplementary Fig. S3), which enabled fluid handling and maintained a constant 5 μL volume in the system.


Label-free biomarker detection from whole blood.

Stern E, Vacic A, Rajan NK, Criscione JM, Park J, Ilic BR, Mooney DJ, Reed MA, Fahmy TM - Nat Nanotechnol (2009)

Schematic of MPC operationa, Primary antibodies to multiple biomarkers, here prostate specific antigen (PSA) and carbohydrate antigen-15.3 (CA15.3), are bound with a photocleavable crosslinker to the MPC. The chip is placed in a plastic housing and a valve (pink) directs fluid flow exiting the chip to either a waste receptacle or the nanosensor chip. b, Whole blood is injected into the chip with the valve set to the waste compartment (black arrow shows the direction of fluid flow) and, if present in the sample, biomarkers bind their cognate antibodies. c, Washing steps follow blood flow and the chip volume (5 μL) is filled with sensing buffer prior to UV irradiation (orange arrows). During UV exposure, the photolabile crosslinker cleaves, releasing the antibody-antigen complexes into solution. d, The valve is set to the nanosensor reservoir (black arrow shows the direction of fluid flow) and the 5 μL volume is transferred, enabling label-free sensing to be performed to determine the presence of specific biomarkers.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2818341&req=5

Figure 1: Schematic of MPC operationa, Primary antibodies to multiple biomarkers, here prostate specific antigen (PSA) and carbohydrate antigen-15.3 (CA15.3), are bound with a photocleavable crosslinker to the MPC. The chip is placed in a plastic housing and a valve (pink) directs fluid flow exiting the chip to either a waste receptacle or the nanosensor chip. b, Whole blood is injected into the chip with the valve set to the waste compartment (black arrow shows the direction of fluid flow) and, if present in the sample, biomarkers bind their cognate antibodies. c, Washing steps follow blood flow and the chip volume (5 μL) is filled with sensing buffer prior to UV irradiation (orange arrows). During UV exposure, the photolabile crosslinker cleaves, releasing the antibody-antigen complexes into solution. d, The valve is set to the nanosensor reservoir (black arrow shows the direction of fluid flow) and the 5 μL volume is transferred, enabling label-free sensing to be performed to determine the presence of specific biomarkers.
Mentions: Figure 1 schematically illustrates the operation of the MPC chip. The avidin-functionalized chip19 (Fig. 1a) is treated with antibodies to any number of specific biomarkers conjugated to biotinylated, photocleavable crosslinkers containing a specific 19-mer DNA sequence (Fig. 2a)20. The MPC geometry was chosen to optimize biomarker binding (Supplementary Fig. S1)14 and chips were fabricated from 4-inch silicon wafers in a one-step photolithographic process (Supplementary Fig. S2). Completed chips (Fig. 2b) were loaded into a custom-machined flow chamber (inset, Fig. 2b and Supplementary Fig. S3), which enabled fluid handling and maintained a constant 5 μL volume in the system.

Bottom Line: However, detecting these biomarkers in physiological fluid samples is difficult because of problems such as biofouling and non-specific binding, and the resulting need to use purified buffers greatly reduces the clinical relevance of these sensors.This two-stage approach isolates the detector from the complex environment of whole blood, and reduces its minimum required sensitivity by effectively pre-concentrating the biomarkers.This study marks the first use of label-free nanosensors with physiological solutions, positioning this technology for rapid translation to clinical settings.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, School of Engineering and Applied Science, Yale University, New Haven, Connecticut 06511, USA.

ABSTRACT
Label-free nanosensors can detect disease markers to provide point-of-care diagnosis that is low-cost, rapid, specific and sensitive. However, detecting these biomarkers in physiological fluid samples is difficult because of problems such as biofouling and non-specific binding, and the resulting need to use purified buffers greatly reduces the clinical relevance of these sensors. Here, we overcome this limitation by using distinct components within the sensor to perform purification and detection. A microfluidic purification chip simultaneously captures multiple biomarkers from blood samples and releases them, after washing, into purified buffer for sensing by a silicon nanoribbon detector. This two-stage approach isolates the detector from the complex environment of whole blood, and reduces its minimum required sensitivity by effectively pre-concentrating the biomarkers. We show specific and quantitative detection of two model cancer antigens from a 10 microl sample of whole blood in less than 20 min. This study marks the first use of label-free nanosensors with physiological solutions, positioning this technology for rapid translation to clinical settings.

Show MeSH
Related in: MedlinePlus