Solute-phase aptamer sensing with aptamer isolation

Abstract

A sensing device for sensing at least one analyte in a sample solution is provided. The device 100 includes a sensor fluid 18 including an aptamer, and the aptamer is configured to measure an analyte included in a sample fluid 14 , the aptamer configured to freely diffuse in the sensor fluid. The device also includes at least one isolation element (e.g., membrane 136 ) retaining the aptamer in the sensor fluid. A method of fabrication of a sensing device for sensing at least one analyte in a sample solution is provided. The method includes defining a volume 130 configured to house a sample fluid with at least one analyte, measuring an analyte included in a sensor fluid that is at least in part measured by an aptamer freely diffusing in the sensor fluid, and retaining the aptamer in the sensor fluid with at least one isolation element. A method of sensing an analyte in a sample fluid is also provided. The method includes bringing an analyte included in a sample fluid into contact with an aptamer included in a sensor fluid, resulting in a change in the electron transfer between a redox tag and an electrode, and measuring the change in electron transfer between the redox tag and the electrode.

What is claimed is: 
     
         1 . A sensing device for sensing at least one analyte comprising:
 a sensor fluid including a plurality of aptamers, one or more aptamers of the plurality of aptamers being configured to sense an analyte included in a sample fluid, the plurality of aptamers configured to freely diffuse in the sensor fluid; and   at least one isolation element retaining the plurality of aptamers in the sensor fluid.   
     
     
         2 . The device of  claim 1 , wherein the isolation element is a membrane. 
     
     
         3 . The device of  claim 1 , wherein the one or more aptamers of the plurality of aptamers are configured to measure the analyte electrochemically and the one or more aptamers each includes at least one redox tag and the device includes at least one electrode. 
     
     
         4 . The device of  claim 1 , wherein the one or more aptamers of the plurality of aptamers are configured to measure the analyte optically and the one or more aptamers includes at least one fluorescent tag, at least one optical source, and at least one optical detector. 
     
     
         5 . The device of  claim 1 , further comprising a reservoir fluid that is in fluidic communication with the sensor fluid. 
     
     
         6 . The device of  claim 5 , wherein a volume of reservoir fluid is at least one of 2×, 10×, 50×, 250× greater than a volume of the sensor fluid. 
     
     
         7 . The device of  claim 5 , wherein a first mass flow of aptamer through the isolation element and a second mass flow of aptamer through a fluidic connection between the sensor fluid and the reservoir fluid, and the first mass flow is at least 2×, 10×, 50×, 250× less than the second mass flow. 
     
     
         8 . The device of  claim 5 , wherein there is a first mass flow of analyte through the isolation element and a second mass flow of analyte through a fluidic connection between the sensor fluid and the reservoir fluid, and the first mass flow is at least 2×, 10×, 50×, 250× greater than the second mass flow. 
     
     
         9 . The device of  claim 7 , wherein a concentration of aptamer in the sensor fluid is within at least 50%, 10%, 2%, 0.4% of the concentration in the reservoir fluid. 
     
     
         10 . The device of  claim 8 , where in the concentration of analyte in the sensor fluid is within at least 50%, 10%, 2%, 0.4% of the concentration in the sample fluid. 
     
     
         11 . The device of  claim 2 , wherein the membrane includes a backing material and the backing material is facing the sample fluid. 
     
     
         12 . The device of  claim 2 , wherein the membrane has a Deff/Δx that is greater than an amount selected from the group consisting of 5 m s −1 ×10 −3 , 0.5 m s −1 ×10 −3 , 0.05 m s −1 ×10 −3 , and 0.005 m s −1 ×10 −3 . 
     
     
         13 . The device of  claim 2 , wherein the membrane has a thickness that is less than an amount selected from the group consisting of ˜100 nm, 1 μm, 10 μm and 100 μm thick. 
     
     
         14 . The device of  claim 2 , wherein the membrane has a change in retentivity/change in molecular weight that is greater than an amount selected from the group consisting of 2×, 5×, 10×, and 20×. 
     
     
         15 . The device of  claim 5 , wherein the device is configured to retain 90% of the initial aptamer concentration in the sensor fluid for a period of time selected from the group consisting of >16 months, >8 months, >4 months, >2 months, >1 month, >2 weeks, and >1 week. 
     
     
         16 . The device of  claim 1 , wherein each aptamer of the plurality of aptamers includes at least an active portion and an inactive portion, the inactive portion comprising at least 50% of the total aptamer molecular weight. 
     
     
         17 . The device of  claim 16 , wherein the inactive portion of the aptamer is rigid by having a secondary structure wherein the aptamer is bound to itself. 
     
     
         18 . The device of  claim 16 , wherein the inactive portion of the aptamer includes at least one permanent fold. 
     
     
         19 . The device of  claim 16 , wherein the aptamer has a molecular weight that is an amount selected from the group consisting of >15 kDa, >30 kDa, or >60 kDa and >120 kDa. 
     
     
         20 . The device of  claim 16 , wherein the active portion of the aptamer has a molecular weight that is an amount selected from the group consisting of <20, <10, and <5 kDa. 
     
     
         21 . The device of  claim 1 , wherein a majority of the plurality of aptamers are bound to a plurality of nanoparticles. 
     
     
         22 . The device of  claim 21 , wherein the nanoparticles are magnetic and the isolation element is a magnet. 
     
     
         23 . The device of  claim 21 , wherein the size of the particles is selected from the group consisting of >1 nm, >3 nm, >10 nm, >30 nm, and >100 nm in diameter. 
     
     
         24 . The device of  claim 1 , wherein the analyte is a small molecule of molecular weight less than 1000 Da. 
     
     
         25 . The device of  claim 1 , wherein the analyte is a protein. 
     
     
         25 . The device of  claim 1 , wherein the sensor fluid includes at least one passivating solute. 
     
     
         26 . The device of  claim 1 , wherein the device is a continuous sensing device. 
     
     
         27 . The device of  claim 1 , wherein the device is a single-use device. 
     
     
         28 . The device of  claim 1 , wherein the device has a lag time, and the lag time to reach 90% of a sensor response is less than at least one of 180 min, 60 min, 20 min, 5 min, 2 min. 
     
     
         29 . The device of  claim 1 , wherein the device further comprises a surface opposite the isolation element, the surface defining a volume housing the sensor fluid and a distance between the surface and the element is at least one of less than 100 μm, 10 μm, 1 μm, 0.1 μm, 0.01 μm. 
     
     
         30 . The device of  claim 3 , wherein the electrode has an area and the membrane has an area, and the electrode area and membrane area have a ratio of <1, <0.1, <0.01 or <0.001. 
     
     
         31 . The device of  claim 3 , wherein a passivating layer is present on the electrode. 
     
     
         32 . The device of  claim 3 , wherein the redox tag is configured to move within the sensor fluid, and the movement of the redox tag results is a change of electron transfer between the redox tag and the electrode. 
     
     
         33 . The device of  claim 1 , wherein the plurality of aptamers comprise a plurality of signaling aptamers and a plurality of anchor aptamers. 
     
     
         34 . The device of  claim 33 , wherein the plurality of redox tags are bound to the signaling aptamers, but are not bound to the anchor aptamers. 
     
     
         35 . The device of  claim 1 , wherein the aptamer is a folded aptamer. 
     
     
         36 . The device of  claim 1 , wherein the aptamer is an unfolded aptamer. 
     
     
         37 . The device of  claim 2 , wherein the membrane has a Deff/Δx of between about 5 m s −1 ×10 −3  and 0.005 m s −1 ×10 −3 . 
     
     
         38 . The device of  claim 2 , wherein the membrane has a Deff/Δx that is at least 0.005 m s −1 ×10 −3 . 
     
     
         39 . The device of  claim 2 , wherein the device is configured to measure 90% of the sample fluid concentration in at least <15 min. 
     
     
         40 . The device of  claim 1 , wherein one or more aptamers of the plurality of aptamers is tagged with a fluorescent optical tag. 
     
     
         41 . The device of  claim 41 , further comprising a fluorescence quencher. 
     
     
         42 . A method of fabricating a sensing device for at least one analyte comprising:
 defining a first volume configured to house a sample fluid with at least one analyte;   defining a second volume configured to house a sensor fluid, the sensor fluid including an analyte that is at least in part measured by an aptamer freely diffusing in the sensor fluid;   retaining the aptamer in the sensor fluid with at least one isolation element.   
     
     
         43 . The method of  claim 42 , wherein the aptamer is coated as a solid layer and the isolation element is coated onto a solid layer including the aptamer. 
     
     
         44 . The method of  claim 43 , wherein spacer material is coated along with the aptamer in the solid layer. 
     
     
         45 . The method of  claim 42 , further comprising fabricating a solid aptamer layer and the isolation element with solvent systems that are orthogonal; and fabricating the isolation element configured to be filled with a solution during use of the device with a sample fluid. 
     
     
         46 . The method of  claim 42 , further comprising defining a reservoir configured to house a reservoir fluid, wherein the sensor fluid is in fluid communication with the reservoir. 
     
     
         47 . The method of  claim 42 , wherein the sensor fluid is in fluid communication with the reservoir via a pore. 
     
     
         48 . The method of  claim 42 , wherein the isolation element is a membrane. 
     
     
         49 . The method of  claim 42  further comprising housing an electrode in the second volume. 
     
     
         50 . The method of  claim 49  further comprising spacing the electrode and the isolation element at least one of less than 100 μm, 10 μm, 1 μm, 0.1 μm, or 0.01 μm apart. 
     
     
         51 . A method of sensing an analyte in a sample solution, the method comprising:
 bringing an analyte included in a sample fluid into contact with an aptamer included in a sensor fluid, the aptamer being freely diffusing in the sensor fluid, the contact of the aptamer with the analyte resulting in a change in the electron transfer between a redox tag and an electrode; and   measuring the change in electron transfer between the redox tag and the electrode.   
     
     
         52 . The method of  claim 51 , wherein the sample fluid is in fluid communication with the sensor fluid, and wherein the sensor fluid is in fluid communication with a reservoir fluid, the method further comprising:
 flowing the aptamer from the sensor fluid to the sample fluid at a first rate; and   flowing the aptamer from the reservoir fluid to the sensor fluid at a second flow rate.   
     
     
         53 . The method of  claim 52 , wherein the first rate is at least 2×, 10×, 50×, or 250× less than the second rate. 
     
     
         54 . The method of  claim 51 , wherein the sample fluid is in fluid communication with the sensor fluid, and wherein the sensor fluid is in fluid communication with a reservoir fluid, the method further comprising:
 flowing the analyte from the sensor fluid to the sample fluid at a first rate; and   flowing the analyte from the sensor fluid to the reservoir fluid at a second flow rate.   
     
     
         55 . The method of  claim 54 , wherein the first rate is at least at least 2×, 10×, 50×, or 250× greater than the second rate. 
     
     
         56 . The method of  claim 51 , wherein the analyte is a protein. 
     
     
         57 . The method of  claim 51 , wherein measuring a change in electron transfer between the redox tag and the electrode has a lag time, and the lag time to reach 90% of a sensor response is less than at least one of 180 minutes, 60 minutes, 20 minutes, 5 minutes, and 2 minutes. 
     
     
         58 . The method of  claim 51 , wherein the redox tag is attached to the aptamer.