WO2022025864A1

WO2022025864A1 - Microfluidic chips with plasmonic sensors

Info

Publication number: WO2022025864A1
Application number: PCT/US2020/043828
Authority: WO
Inventors: Fausto D'APUZZO; Raghuvir N. SENGUPTA; Albert Liu; Viktor Shkolnikov
Original assignee: Hewlett-Packard Development Company, L.P.
Priority date: 2020-07-28
Filing date: 2020-07-28
Publication date: 2022-02-03
Also published as: TW202219276A

Abstract

In example implementations, a microfluidic chip is provided. The microfluidic chip includes a first channel, a second channel, and a plasmonic sensor. The first channel includes a first antibiotic chamber and a first filter chamber. The first antibiotic chamber includes a first media and a first antibiotic that is rehydrated by a bacterial suspension and the first filter chamber is to filter bacteria from first metabolites released from the bacteria in response to incubation with a buffer solution. The second channel includes a second antibiotic chamber and a second filter chamber. The second antibiotic chamber includes a second media and a second antibiotic that is rehydrated by the bacterial suspension and the second filter chamber is to filter the bacteria from second metabolites released from the bacteria in response to incubation with the buffer solution. The first channel and the second channel pass over the plasmonic sensor. The plasmonic sensor is to measure an amount of the first metabolites and the second metabolites.

Description

MICROFLUIDIC CHIPS WITH PLASMONIC SENSORS

BACKGROUND

[0001] Antibiotic susceptibility testing (AST) may be used to analyze the effectiveness of antibiotics for a particular strain of bacteria. AST can be used for proper and effective prescription of antibiotics and dosages that are administered to patients. The AST can analyze the effectiveness of antibiotics by measuring an amount of bacteria in solution with antibiotics. For example, bacteria may be mixed with a nutrient media that allows the bacteria to grow. The nutrient media may be replaced with water that causes the bacteria to undergo metabolic changes in response to the absence of nutrients. To persist in this environment, the bacterial cells may begin a series of catabolic reactions and the byproducts of these processes may be purine metabolites that are released. The presence of the purine metabolites may be analyzed to determine the particular strain of bacteria.

[0002] In some examples, bacteria may be incubated with nutrient media mixed with antibiotics and subsequently transferred to water. Changes in the amount of the purine metabolites in response to antibiotics may be analyzed to determine the effectiveness of the antibiotic.

[0003] Some AST methods have a turn-around time that can be several hours. For example, turn-around times can be 12-24 hours. These methodologies may also include bacterial culture and isolation steps that precede the AST that can have an additional processing time of 18-24 hours.

BRIEF DESCRIPTION OF THE DRAWINGS

[0004] FIG. 1 illustrates a block diagram of an example microfluidic chip with a plasmonic sensor of the present disclosure;

[0005] FIG. 2 illustrates a top view of a block diagram of another example of the microfluidic chip with the plasmonic sensor of the present disclosure;

[0006] FIG. 3 illustrates a cross-sectional view of the example microfluidic chip with the plasmonic sensor illustrated in FIG. 2 of the present disclosure; [0007] FIG. 4 illustrates an example workflow diagram for performing antibiotic susceptibility testing (AST) with the microfluidic chip with the plasmonic sensor illustrated in FIG. 2 of the present disclosure;

[0008] FIG. 5 illustrates a top view of a block diagram of another example of the microfluidic chip with the plasmonic sensor of the present disclosure;

[0009] FIG. 6 illustrates an example workflow diagram for performing AST with the microfluidic chip with the plasmonic sensor illustrated in FIG. 5 of the present disclosure;

[0010] FIG. 7 illustrates a top view of an example of how the microfluidic channels can be routed over the plasmonic sensor of the present disclosure; and

[0011] FIG. 8 illustrates an example flowchart for a method of performing AST with a microfluidic chip with a plasmonic sensor of the present disclosure.

DETAILED DESCRIPTION

[0012] Examples described herein provide a microfluidic chip with a plasmonic sensor to perform antibiotic susceptibility testing (AST) of the present disclosure. As noted above, AST may be used to analyze the effectiveness of antibiotics for a particular strain of bacteria. However, current methodologies may have a turn-around time of several hours (e.g., 12-24 hours).

[0013] However, due to these long turn-around times, most patients are prescribed a broad spectrum of antibiotics when there is suspicion of an infection. These broad-spectrum prescriptions sometimes have no effect at all because of the spread of antibiotic-resistant bacterial strains. In these situations, patient conditions can degrade rapidly and chance of life-threatening complications can increase by the hours.

[0014] The present disclosure provides a microfluidic chip with a plasmonic sensor that can reduce the turn-around time for AST to several minutes to a few hours (e.g., less than 2 hours). The microfluidic chip of the present disclosure is relatively low cost, with high accuracy that can be scaled for large distribution.

In addition, the microfluidic chip with the plasmonic sensor can analyze multiple different concentrations or types of antibiotics for a bacterial strain within a single microfluidic chip. Thus, the present disclosure can reduce the turn around time for AST that may allow a specific antibiotic with a proper dosage to be prescribed to a patient with an infection.

[0015] FIG. 1 illustrates a block diagram of an example microfluidic chip with a plasmonic sensor 100 (also referred to herein as a chip 100) of the present disclosure. In an example, the chip 100 may include an inlet 102, channels

104₁ and 104₂, and a plasmonic sensor 120. Although two channels 104i and

104₂ are illustrated in FIG. 1 , it should be noted that any number of channels 104 may be deployed. Examples of how the chip 100 may be deployed or manufactured are discussed in further detail below with reference to FIGs. 2-6. Although a single inlet 102 is illustrated in FIG. 1 , it should be noted that separate inlets 102 for the channels 104 may also be deployed.

[0016] In an example, the channel 104i may include a first antibiotic chamber 106i and a first filter chamber 110i . Although shown as separate chambers, in some implementations the first antibiotic chamber 106i and the first filter chamber 110i may be combined as a single chamber. The first antibiotic chamber 106i may include a first antibiotic or a first dosage of an antibiotic and a media 108i (hereinafter also referred to together as simply media 108i). The media 108i may be a lyophilized mixture of the antibiotic and the media. In an example the media 108i may include nutrients that help the bacteria to grow, as discussed in further details below. Examples of the media 108i may include Luria-Bertani (LB) broth, tryptic soy broth, nutrient broth, and the like.

[0017] In an example, the media 108i may be dispensed into the first antibiotic chamber 106i when the chip 100 is fabricated. The media 108i may be prepared during manufacturing of the chip 100 and dispensed via a freeze- dry method or an inkjet printing method. [0018] The first filter chamber 110i may include a filter 112i. The filter 112i may be used to filter out metabolites 116 generated by bacteria 114 in the first filter chamber 110i . The metabolites 116 may be filtered out of the first filter chamber 110i through a first outlet 122i and over the plasmonic sensor 120. [0019] In an example, the channel 104₂ may include a second antibiotic chamber 106₂ and a second filter chamber 110₂. Although shown as separate chambers, in some implementations the second antibiotic chamber IO6₂ and the second filter chamber 110₂ may be combined as a single chamber. The second antibiotic chamber IO6₂ may include a second antibiotic or a second dosage of an antibiotic and a media IO8₂ (hereinafter also referred to together as simply media 108₂). Similar to the media 108₁ , the media 108₂ may be a lyophilized mixture of the antibiotic and the media. In an example the media IO8₂ may include nutrients that help the bacteria to grow, as discussed in further details below. Examples of the media IO8₂ may include Luria-Bertani (LB) broth, tryptic soy broth, nutrient broth, and the like.

[0020] In an example, the media IO8₂ may be dispensed into the second antibiotic chamber 106₂ when the chip 100 is fabricated. The media 108₂ may be prepared during manufacturing of the chip 100 and dispensed via a freeze- dry method or an inkjet printing method.

[0021] The second filter chamber 110₂ may include a filter 112₂. The filter 112₂ may be used to filter out metabolites 116 generated by bacteria 114 in the second filter chamber 110₂. The metabolites 116 may be filtered out of the second filter chamber 110₂ through a second outlet 122₂ and over the plasmonic sensor 120.

[0022] In an example, a bacterial suspension may be introduced to the chip 100 via the inlet 102. The bacterial suspension may be controlled to flow into the first antibiotic chamber IO6₁ and the second antibiotic chamber IO6₂. The bacterial suspension may rehydrate the media IO8₁ and IO8₂, respectively.

After an incubation period (e.g., several minutes), the mixture in the first antibiotic chamber IO6₁ and the second antibiotic chamber IO6₂ may be moved into the first filter chamber 110i and the second filter chamber 110₂.

[0023] In an example, a buffer solution may be introduced to the chip 100 via the inlet 102 (or a separate inlet, as noted above). The buffer solution may be water or another fluid to rinse the bacteria in the bacterial suspension and remove the media and antibiotic 108i and IO82, as well as metabolites released by the micro-organism in the first filter chamber 1101 and the second filter chamber 11O2.

[0024] With the media I O81 and I O82 removed, remaining bacteria 114 in the first filter chamber 110i and the second filter chamber 1102 may be allowed to incubate to induce a stress response. The bacteria 114 may release stress- induced metabolites 116 due to the stress response. The metabolites 116 may be flowed through the filters 112i and 112₂, respectively, and towards the plasmonic sensor 120.

[0025] In an example, the filters 112i and 112₂ may be porous filter membranes. In another example, the filters 112i and 112₂ may include physical structures to allow the metabolites 116 to pass through, while blocking the bacteria 114. For example, the structures may include vertical posts. In an example, different sized microbeads with decreasing diameters may be loaded between the posts to achieve a desired porosity.

[0026] The plasmonic sensor 120 may be used with an optical sensor to measure an amount of metabolites 116 in the outlet channels 122i and 122₂. In an example, the plasmonic sensor 120 may be a surface enhanced Raman spectroscopy (SERS) sensor, a surface enhanced infrared absorption (SEIRA) sensor, a surface enhanced fluorescence (SEF) sensor, surface enhanced luminescence (SEL), and the like. A light source may emit light onto the outlet channels 122i and 122₂ to induce detection. The light rays or beams may be scattered by the plasmonic sensor 120 and ready by an optical detector or sensor. The scattered light rays may be converted into an image or graph by the optical detector that may correspond to an amount of the metabolites 116 in the outlets 122i and 1222.

[0027] The different measurements may determine which antibiotic or dosage of antibiotic in the first antibiotic chamber I O61 and the second antibiotic chamber I O62 is more effective against the bacteria 114. As a result, a proper antibiotic and a proper dosage of the antibiotic may be prescribed based on analysis performed using the chip 100.

[0028] In an example, the chip 100 may also include a temperature control (not shown). The temperature control may be used to control the temperature of the antibiotic chambers 106i and 106₂ and/or the filter chambers 110i and 11O₂. The temperature control may help to mimic the natural habitat of the bacteria 114 or accelerate the production of the metabolites 116 from the stress- induced response of the bacteria 114.

[0029] In an example, temperature control may be external. For example, the chip 100 may be kept in a temperature controlled space, such as an incubator. In an example, the temperature control may be part of the chip. For example, the temperature control may include a resistor and a temperature sensor (e.g., a thermistor) on the substrate of the chip 100. In an example, the temperature control may be a thin film resistor.

[0030] FIG. 2 illustrates an example of a microfluidic chip with a plasmonic sensor 200 (also referred to herein as a chip 200). In an example, the chip 200 may be fabricated as a multi-layered chip with an integrated plasmonic sensor 220. FIG. 2 illustrates a top view of the chip 200.

[0031] In an example, the chip 200 may include an inlet 201 and an inlet 202. The inlet 201 may be used to introduce a bacterial suspension. The bacterial suspension may include a particular strain of bacteria that is to be analyzed. The inlet 202 may be used to introduce a buffer solution.

[0032] The chip 200 may include a plurality of channels 204i to 204_n (hereinafter also referred to individually as a channel 204 or collectively as channels 204). Each channel 204 may include a respective antibiotic chamber 206 and a respective filter chamber 210. The antibiotic chamber 206 may include a lyophilized mixture of an antibiotic and a media 208. The media 208 may be introduced into the antibiotic chamber 206, similar to the media 108₁ and IO8₂ in the chip 100, as discussed above.

[0033] The filter chamber 210 may include a filter 212. The filter 212 may be a porous membrane or a physical structure (e.g., a post). The filter 212 may allow metabolites 216 to pass through, while blocking bacteria 214. The metabolites 216 may flow through the channel over the plasmonic sensor 220. [0034] In an example, the filter 212 may include a second lyophilized mixture 213. The lyophilized mixture may include accelerants as well as a calibration molecule. In an example, the calibration molecule can be used as an internal standard for more quantitative SERS measurements.

[0035] In an example, the accelerants may speed up the stress response of the bacteria 214. The accelerants may include phosphates that can cause the bacteria 214 to release purine metabolites instantaneously or within a few minutes of incubation. This may also help reduce the turn-around time for performing AST in the chip 200. In an example, the accelerants may include disphosphate, phosphoribosyltransferases, nucleoside monophosphates, and the like.

[0036] In an example, each one of the channels 204 may include the antibiotic chamber 206 and the filter chamber 210. Each antibiotic chamber 206 may include different antibiotics or different dosages of the same antibiotic. In an example, one of the channels 204 may include a reporter molecule for in-situ calibration of the optical sensor 224. Each channel 204 may run across the plasmonic sensor 220.

[0037] In an example, the chip 200 may also include capillary breaks 218. The capillary breaks 218 may allow a gas or air to be passed through a channel (shown in the FIG. 3) to control the flow of fluids within the channels 204 of the chip 200. The chip 200 may also include an outlet 222 to allow fluids to exit the chip 200 via vias 230 located between layers of the chip 200.

[0038] In an example, the chip 200 may also include a temperature control (not shown). The temperature control may be used to control the temperature of the antibiotic chambers 206 and/or the filter chambers 210. The temperature control may help to mimic the natural habitat of the bacteria 214 or accelerate the production of the metabolites 216 from the stress-induced response of the bacteria 214.

[0039] In an example, temperature control may be external. For example, the chip 200 may be kept in a temperature controlled space, such as an incubator. In an example, the temperature control may be part of the chip. For example, the temperature control may include a resistor and a temperature sensor (e.g., a thermistor) on the substrate of the chip 200. In an example, the temperature control may be a thin film resistor.

[0040] FIG. 3 illustrates an example cross-sectional side view of the chip 200. As can be seen in FIG. 3, the chip 200 may include a first layer 250, a second layer 252, and a third layer 254. The first layer 250 may include a substrate 256 which the plasmonic sensor 220 is fabricated upon. The substrate 256 may be a semiconductor (e.g., a silicon substrate). The remaining layers may be formed from a photo-definable polymer (e.g., SU8).

The channels 204, antibiotic chambers 206, filter chambers 210, vias 230, and the outlet 222 may be formed via a photo-lithography process that defines and etches the features out of the photo-definable polymer.

[0041] In an example, a portion of the first layer 250 may include an optically clear layer 258. The optically clear layer 258 may provide a window over the plasmonic sensor 220 that allows light beams to reach the channels 204 over the plasmonic sensor 220.

[0042] In an example, the filter 212 may be disposed between the second layer 252 and the first layer 250. In an example, the filter 212 may be selectively deployed at locations where the channel 204 is open between the first layer 250 and the second layer 252. Said another way, the filter 212 may be included in portions of the second layer 252 that include the filter chamber 210.

[0043] In an example, the capillary breaks 218 may be formed as a membrane layer located between the second layer 252 and the third layer 254. The fluid may flow across a channel 232 in the third layer 254 and across the membrane to create a vacuum. The fluid may be controlled to flow over different portions of the membrane at certain capillary breaks 218 to control the flow of fluid within the channels 204.

[0044] In an example, the membrane may include a nano-porous hydrophobic membrane that prevents liquids from flowing through, but allows air or gasses to flow through. The membrane may have pores that have an average diameter of approximately 200 to 400 nanometers (nm) and be fabricated from polytetrafluoroethylene (PTFE). Other example materials for the membrane may include poly(propylene), poly(ethylene), and the like.

[0045] In an example, an optical sensor 224 and a light source 226 may be used to perform the Raman measurements on the chip 200. In one example, the optical sensor 224 may be a Raman spectrometer or a charge coupled device (CCD) detector that generates a CCD image. The light source 226 may be a laser light source. However, the light source 226 may have a length that is equal to the length of the channel 204 that runs across the plasmonic sensor 220. As a result, measurements can be taken from across the entire portion of the channel 204 that runs across the plasmonic sensor 220. In an example, the light source 226 may illuminate each channel 204 across the plasmonic sensor 220 simultaneously. In an example, multiple light sources 226 may be deployed, or a single light source 226 with an optical grating may be deployed. [0046] In an example, the optical sensor 224 may include lenses, filters, diffraction gratings, and other devices (not shown) to focus the incoming light scattered by the plasmonic sensor 220 onto a detector array. In an example, the optical sensor 224 may divide the incoming light into different channels, each of which are sent to a different sensor within the optical sensor 224, providing multi-spectral analysis of the light scattered by the plasmonic sensor 220. The optical sensor 224 may perform brightfield, dark-field, fluorescence, hyperspectral, and other optical analyses.

[0047] FIG. 4 illustrates an example workflow diagram for a method 400 for performing AST with the chip 200 illustrated in FIGs. 2 and 3. At block 402, the method 400 loads a bacterial suspension via the inlet 201. The bacterial suspension may include a particular bacterial strain that is to be examined or analyzed for an effective antibiotic or dosage of antibiotic. The bacterial suspension may be delivered to the antibiotic chamber 206. For example, a fluid may flow across the capillary break 218 of the antibiotic chamber 206 to create a vacuum that pulls the bacterial suspension into the antibiotic chamber 206. Excess bacterial suspension may be removed through the via 230 and the outlet 222.

[0048] At block 404, air 260 may be forced through the channels 204 to purge the loading channels. The air 260 may remove any residual bacterial suspension that may remain in the channels 204.

[0049] At block 406, the bacterial suspension may rehydrate the media 208 and be allowed to incubate for a predefined time period (e.g., several minutes). The nutrients in the media 208 may allow the bacteria to grow and the antibiotics in the media 208 may kill the bacteria. The effectiveness of the antibiotic may be measured based on an amount of metabolites that are released and measured, as discussed in further details below.

[0050] In an example, one of the channels 204 may include a control. Each antibiotic chamber 206 may include a different antibiotic or different dosage of antibiotic. Thus, the chip 200 may allow simultaneous analysis of different antibiotics or dosages against a particular bacterial strain within a relative short turn-around time.

[0051] At block 408, the grown bacteria may be pulled into the filter chamber 210. For example, a fluid may flow across the capillary break 218 of the filter chamber 210 to create a vacuum that pulls the grown bacteria from the antibiotic chamber 206 into the filter chamber 210. The filter chamber 210 may include a lyophilized mixture 213. As described above, the lyophilized mixture 213 may include a calibration molecule and/or an accelerant.

[0052] At block 410, a buffer solution may be introduced via the inlet 202. The buffer solution may be water, a solvent, or any other liquid that can be used to wash the residual media mixed with the bacteria 214 in the filter chamber 210. Again, the capillary break 218 in the filter chamber 210 may be used to pull in the buffer solution. The excess buffer solution may be removed through the via 230 and the outlet 222.

[0053] At block 412, the bacteria 214 may be allowed to incubate without the nutrients in the media. The incubation may induce a stress response that causes the bacteria 214 to release metabolites 216. In an example, the accelerants from the lyophilized mixture 213 may interact with the bacteria 214 in the filter chamber 210 to accelerate the release of metabolites 216 instantaneously or within a few minutes.

[0054] At block 414, the metabolites 216 are transferred through the filter 212 and into the channel 204 over the plasmonic sensor 220. For example, a fluid may flow across the capillary breaks 218 across the ends of the channel 204 to pull the metabolites 216 through the filter 212 and over the plasmonic sensor 220. As discussed above, the filter 212 may be a porous filter membrane or structures (e.g., posts) that allow the metabolites 216 to pass through the filter 212, but prevent the bacteria 214 from passing through the filter 212.

[0055] At block 416, air may be delivered through the inlet 202 to dry the channels 204. The channels may be dried from any residual liquid and the remaining bacteria 214 may be removed via the outlet 222.

[0056] At block 418, a Raman measurement may be performed. As described above, a light source may direct light beams over the plasmonic sensor 220. The light may be scattered by the plasmonic sensor 220 and detected by the optical sensor 224 illustrated in FIGs. 2 and 3.

[0057] As discussed above, the chip 200 may allow different antibiotics and/or dosages to be analyzed. Based on the measurements, the most effective antibiotic or dosage may be selected to treat an infection with the particular bacterial strain. The turn-around time for analysis with the chip 200 may be several minutes to a few hours. In addition, the relatively low cost to manufacture the chip 200 may allow the chip 200 to be scaled for large volume production and manufacturing.

[0058] FIG. 5 illustrates a top view of a block diagram of another example of a microfluidic chip with a plasmonic sensor 500 (hereinafter also referred to as a chip 500). In an example, the chip 500 may be fabricated along parallel channels 504i to 504_n (hereinafter also referred to individually as a channel 504 or collectively as channels 504) on a single layer. For example, the channels 504 may lie on a single horizontal plane across a substrate.

[0059] In an example, the chip 500 may include an inlet 501 , a plurality of channels 504i to 504_n, and an outlet 530. Although a single inlet 501 is illustrated in FIG. 5, it should be noted that multiple inlets 501 may be deployed. [0060] Each channel 504 may include a respective antibiotic chamber 506i to 506_n and a respective filter chamber 510i to 510_n. Each antibiotic chamber 506i to 506_n may include a respective antibiotic and media 508i to 508_n. Each filter chamber 510i to 510_n may include a respective filter 512i to 512_n. The filters 512 may be porous filter membranes or structures (e.g., posts), as described above.

[0061] Each filter chamber 510i to 510_n may branch off into two separate outlet channels 518 and 522. The outlet channel 518 may include a plasmonic sensor 520. For example, the filter chamber 510i may include outlet channels 518i and 522i. The outlet channel 518i may include a plasmonic sensor 520i. The filter chamber 5102 may include outlet channels 5182 and 522₂. The outlet channel 5182 may include a plasmonic sensor 520₂. The filter chamber 510_n may include outlet channels 518_n and 522_n. The outlet channel 518_n may include a plasmonic sensor 520_n.

[0062] In an example, the chip 500 may include valves 504i to 504_n, valves 509i to 509_n, valves 514i to 514_n, and valves 5161 to 516_n. The valves 504,

509, 514, and 516 may be microfluidic valves that are operated mechanically or electromechanically. The valves 504, 509, 514, and 516 may be used to control flow of liquids or fluids through the chip 500 and through respective channels 502.

[0063] In an example, the chip 500 may be used together with a light source and optical sensor (not shown), similar to the light source 226 and the optical source 224, described above. The light source 224 may emit light onto each plasmonic sensor 520i to 520_n and the response may be detected and measured by the optical sensor.

[0064] FIG. 6 illustrates an example workflow diagram of a method 600 for performing AST in the chip 500 of the present disclosure. At block 602 a bacterial suspension may be loaded via the inlet 501 . The bacterial suspension may include a particular bacterial strain that is to be examined or analyzed for an effective antibiotic or dosage of antibiotic.

[0065] In an example, the valves 504 may be opened, while the valves 509 remain closed. The bacterial suspension may be delivered to the antibiotic chambers 506. The antibiotic chambers 506 may include a mixture of an antibiotic and a media 508. Each antibiotic chamber 506i to 506_n may include a different antibiotic or a different dosage of the same antibiotic. In an example, one of the antibiotic chambers 506i to 506_n may include a control.

[0066] At block 604, the valves 504 may be closed. The bacterial suspension that includes bacteria 532 may rehydrate the media 508. The media 508 may include nutrients that may allow the bacteria 532 to grow. The antibiotic may kill the bacteria 532. The effectiveness of the antibiotic or a particular dosage of antibiotic may be determined based on the amount of metabolites that are released, which may correlate to an amount of bacteria that remains, as discussed in further details below. The bacteria 532 may be allowed to incubate for a predetermined amount of time (e.g., several minutes to several hours).

[0067] At block 606, valves 504, 509, and 514 may be opened and the valve 516 may be closed. A buffer solution may be delivered via the inlet 501. The buffer solution may push the bacteria 532 and media 508 into the filter chamber 510. The buffer solution may rinse the bacteria 532 to remove the media 508 through the valve 514, through the outlet channel 522, out via the outlet 530. [0068] At block 608, the valves 514 and 516 may be closed to allow the bacteria 532 to incubate in the filter chamber 510 for a predefined amount of time (e.g., several minutes). Without the nutrients in the media 508, the bacteria 532 may undergo a stress response. The stress response may cause the bacteria 532 to release metabolites 534. In an example, an accelerant may be added to the bacteria 532 to accelerate the release of the metabolites 534.

[0069] At block 610, the valve 516 may be opened to allow the metabolites 534 to flow through the channel 518. The metabolites may flow over the respective plasmonic sensor 520 in the channel 518. In an example, the valve 516 may be closed and the valve 514 may be opened to flush the remaining bacteria 532 through the valve 514 and out via the channels 520.

[0070] In an example, the valve 514 may be closed and the valve 516 may be opened to dry the outlet channel 520. For example, air may be blown through the chip 500 to dry the outlet channels 520.

[0071] At block 612, a laser light (e.g., the laser light source 226) may emit light beams or rays over the plasmonic sensors 520i to 520_n. The light scattered by the plasmonic sensors 520 may be read by an optical sensor. The optical sensor may measure an amount of metabolites 536 based on the scattered light signals. The amount of metabolites 536 may be correlated to an amount of bacteria, which then can be used to measure the effectiveness of the antibiotic or dosage within a particular antibiotic chamber 506. The most effective antibiotic or dosage of antibiotic may be selected or prescribed to treat an infection caused by the bacteria 532.

[0072] As discussed above, the chip 500 may allow different antibiotics and/or dosages to be analyzed. Based on the measurements, the most effective antibiotic or dosage may be selected to treat an infection with the particular bacterial strain. The turn-around time for analysis with the chip 500 may be several minutes to a few hours. In addition, the relatively low cost to manufacture the chip 500 may allow the chip 500 to be scaled for large volume production and manufacturing.

[0073] In an example, the chip 500 may also include a temperature control (not shown). The temperature control may be used to control the temperature of the antibiotic chambers 506 and/or the filter chambers 510. The temperature control may help to mimic the natural habitat of the bacteria 532 or accelerate the production of the metabolites 534 from the stress-induced response of the bacteria 532.

[0074] In an example, temperature control may be external. For example, the chip 500 may be kept in a temperature controlled space, such as an incubator. In an example, the temperature control may be part of the chip. For example, the temperature control may include a resistor and a temperature sensor (e.g., a thermistor) on the substrate of the chip 500. In an example, the temperature control may be a thin film resistor.

[0075] FIG. 7 illustrates a top view of an example of how microfluidic channels 702, 704, and 706 may be routed over a plasmonic sensor 720. The channels 702, 704, and 706 may be part of the chip 100, 200, or 500, illustrated in FIGs. 1-6, and discussed above. The plasmonic sensor 720 may be similar to the plasmonic sensor 120, 220, and 520, illustrated in FIGs. 1-6, and discussed above.

[0076] In an example, the channels 702, 704, and 706 may be routed over the plasmonic sensor 720 several times. This may provide an average reading by the optical sensor and provide a more accurate measurement. In an example, each portion of the channels 702, 704, and 706 that goes over the plasmonic sensor 720 may be a straight run. In an example, the channels 702, 704, and 706 may be routed over the plasmonic sensor 720 in a serpentine fashion to achieve multiple passes over the plasmonic sensor 720.

[0077] As can be seen in FIG. 7, the curves or bends may occur outside of the active area of the plasmonic sensor 720. The straight runs may occur over the plasmonic sensor 720. The straight runs may allow a light source that emits light in a line to be used to measure the entire portion of the channels 702, 704, and 706 that located over the plasmonic sensor 720.

[0078] The several lines of light may be emitted to measure each portion of each channel 702, 704, and 706 over the plasmonic sensor 720 simultaneously. For example, in FIG. 7, there are nine lines that cross over the plasmonic sensor 720. The light source may emit nine separate lines of light (e.g., using optical grating or different light sources) to measure all portions of the channels 702, 704, and 706 that cover the plasmonic sensor 720 simultaneously.

[0079] It should be noted that the serpentine pattern illustrated in FIG. 7 is one example pattern. Other patterns may be deployed to achieve the same result. In addition, although three channels are illustrated in FIG. 7, any number of channels may be deployed.

[0080] FIG. 8 illustrates a flowchart for a method 800 for method of performing AST with a microfluidic chip with a plasmonic sensor of the present disclosure. In an example, the method 800 may be performed by the microfluidic chip 100 illustrated in FIG. 1 , the microfluidic chip 200 illustrated in FIGs. 2 and 3, or the microfluidic chip 500 illustrated in FIG. 5, and described above.

[0081] At block 802, the method 800 begins. At block 804, the method 800 rehydrates a first media and a first antibiotic in a first antibiotic chamber with a bacterial suspension to form a first mixture for an incubation period. The first media and the first antibiotic may be a lyophilized mixture that is premixed into the first antibiotic chamber. The bacterial suspension may include a particular strain of bacteria that is to be analyzed (e.g., to treat an infection caused by the strain of bacteria in a patient). The first media may include nutrients that allow the bacteria to grow within the first antibiotic chamber, while being killed by the first antibiotic.

[0082] At block 806, the method 800 rehydrates a second media and a second antibiotic in a second antibiotic chamber with the bacterial suspension to form a second mixture for the incubation period. The second media and the second antibiotic may be a lyophilized mixture that is premixed into the second antibiotic chamber. The bacterial suspension may include a particular strain of bacteria that is to be analyzed (e.g., to treat an infection caused by the strain of bacteria in a patient). The second media may include nutrients that allow the bacteria to grow within the second antibiotic chamber, while being killed by the second antibiotic. In an example, the first antibiotic and the second antibiotic may be different antibiotics or may include different dosages of the same antibiotic.

[0083] At block 808, the method 800 traps the first mixture in a first filter chamber and the second mixture in a second filter chamber. After the bacterial suspension is allowed to grow in the first antibiotic chamber and the second antibiotic chamber, the mixtures may be moved to the first and second filter chambers.

[0084] At block 810, the method 800 washes the first mixture and the second mixture with a buffer solution to induce a stress response from bacteria to release a first metabolite in the first filter chamber and a second metabolite in the second filter chamber. Without the media, the bacteria remaining in the first filter chamber and the second filter chamber may undergo a stress response. The stress response may cause the bacteria to release metabolites.

[0085] In an example, an accelerant may be added to the bacteria. The accelerant may increase the rate at which the bacteria releases sthe metabolites. The metabolites may be measured to determine an amount of bacteria remaining in the first mixture and the second mixture. The remaining amount of bacteria may determine the effectiveness of the first antibiotic and the second antibiotic. [0086] At block 812, the method 800 transfers the first metabolite and the second metabolite over a plasmonic sensor to perform a Raman measurement. For example, the first filter chamber and the second filter chamber may include a filter that allows the metabolites to pass, while blocking the bacteria. The metabolites may flow through the channel and across the plasmonic sensor.

The channels may be dried before performing the Raman measurement. The Raman measurement may determine an amount of the metabolites in each channel. As noted above, the measurement of the metabolites can be used to determine the effectiveness of the first antibiotic and the second antibiotic.

[0087] At block 814, the method 800 generates a recommendation for the first antibiotic or the second antibiotic based on the Raman measurement. For example, a processor within an optical sensor may compare the Raman measurement within each channel. The channel with the lowest amount of bacteria may determine the most effective antibiotic or most effective dosage of the antibiotic. The most effective dosage of antibiotic may be recommended. [0088] Although the method 800 was described with two channels, it should be noted that method 800 may be performed for more than two channels. In addition, one of the channels may include a control (e.g., no antibiotics). At block 816, the method 800 ends.

[0089] It will be appreciated that variants of the above-disclosed and other features and functions, or alternatives thereof, may be combined into many other different systems or applications. Various presently unforeseen or unanticipated alternatives, modifications, variations, or improvements therein may be subsequently made by those skilled in the art which are also intended to be encompassed by the following claims.

Claims

1. A microfluidic chip, comprising: a first channel comprising a first antibiotic chamber and a first filter chamber, wherein the first antibiotic chamber includes a first media and a first antibiotic that is rehydrated by a bacterial suspension, and the first filter chamber is to filter bacteria from first metabolites released from the bacteria in response to incubation with a buffer solution; a second channel comprising a second antibiotic chamber and a second filter chamber, wherein the second antibiotic chamber includes a second media and a second antibiotic that is rehydrated by the bacterial suspension, and the second filter chamber is to filter the bacteria from second metabolites released from the bacteria in response to incubation with the buffer solution; and a plasmonic sensor, wherein the first channel and the second channel pass over the plasmonic sensor, wherein the plasmonic sensor is to measure an amount of the first metabolites and the second metabolites.

2. The microfluidic chip of claim 1 , further comprising: a first inlet to provide the bacterial suspension; and a second inlet to provide the buffer solution.

3. The microfluidic chip of claim 1 , wherein the first filter chamber and the second filter chamber each comprises a filter.

4. The microfluidic chip of claim 1 , further comprising: a multi-layer fluidic chip, comprising: a first layer including a channel to provide a fluid to control flow of the bacterial suspension in the first channel and the second channel; a second layer including the first channel, the first antibiotic chamber, the first filter chamber, the second channel, the second antibiotic chamber, and the second filter chamber; a filter layer coupled to an opening of the first filter chamber and the second filter chamber; and a third layer including the plasmonic sensor and an optically clear window over the plasmonic sensor.

5. The microfluidic chip of claim 4, further comprising: a capillary break membrane layer located between the first layer and the second layer.

6. The microfluidic chip of claim 1 , wherein the first channel and the second channel are arranged in parallel on a common plane.

7. The microfluidic chip of claim 6, wherein the first filter chamber and the second filter chamber each comprises a first valve coupled to a first outlet channel and a second valve coupled to a second outlet channel that includes the plasmonic sensor.

8. The microfluidic chip of claim 1 , wherein the first antibiotic chamber and the first filter chamber comprise a single chamber.

9. The microfluidic chip of claim 1 , wherein the first channel and the second channel are arranged in a serpentine shape to cross over the plasmonic sensor multiple times over different locations of the plasmonic sensor.

10. A microfluidic chip, comprising: an inlet to provide a bacterial suspension and a buffer solution; a plurality of channels, wherein each one of the plurality of channels includes a respective media chamber and a respective filter chamber, wherein the plurality of channels includes a different antibiotic and a media in the respective media chambers, wherein the different antibiotic and the media in the media chambers is rehydrated by the bacterial suspension; a flow control to move a mixture of the media, the different antibiotic, and the bacterial suspension to the respective filter chamber, wherein the buffer solution is to remove the media from the mixture to cause bacteria to release metabolites; and a plasmonic sensor to measure an amount of metabolites in the plurality of channels, wherein the metabolites are filtered from the mixture via a filter in the respective filter chamber.

11. The microfluidic chip of claim 10, wherein one of the plurality of channels includes a control sample.

12. The microfluidic chip of claim 10, wherein the plasmonic sensor comprises a surface-enhanced Raman spectroscopy (SERS) sensor.

13. A method, comprising: rehydrating a first media and a first antibiotic in a first antibiotic chamber with a bacterial suspension to form a first mixture for an incubation period; rehydrating a second media and a second antibiotic in a second antibiotic chamber with the bacterial suspension to form a second mixture for the incubation period; trapping the first mixture in a first filter chamber and the second mixture in a second filter chamber; washing the first mixture and the second mixture with a buffer solution to induce a stress response from bacteria to release a first metabolite in the first filter chamber and a second metabolite in the second filter chamber; transferring the first metabolite and the second metabolite over a plasmonic sensor to perform a Raman measurement; and generating a recommendation for the first antibiotic or the second antibiotic based on the Raman measurement.

14. The method of claim 13, further comprising: mixing an accelerant with the first mixture in the first filter chamber and the second mixture in the second filter chamber.

15. The method of claim 14, wherein the accelerant comprises a phosphoribosyltransferase or a nucleoside monophosphate.