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Highly sensitive continuous flow microfluidic chip sensor with integrated BI/SB thermopile for biochemical applications.


Measuring biochemical process has showed increasing importance with the development of chip calorimeters [1]. Chip calorimetry is generally achieved by integrating the calorimetric detection using sensors such as thermistors and thermopiles with the micro flow channels for sample handling. Several groups have shown the advantages of chip calorimeters for the analysis of biochemical process [2], [3]. Often, chip calorimeters are realized using MEMS fabrication to achieve high heat power sensitivities. MEMS techniques are highly complex, time consuming, expensive and require sophisticated equipment. To monitor biochemical process such as metabolic activity of cells and binding event measurements of bio molecules, the heat power sensitivities of the chip calorimeters must be in the range of nW to pW [4]-[6]. Rapid prototyping has shown great promise in realizing inexpensive and easy to fabricate fluidic chips with micrometer features enabling lab-on-chip analysis [7]. Thermopile sensors generally used in chip calorimeters need extreme reference temperature control. Common mode rejection of thermal signals by the thermopile sensors was explored recently by a few groups [8], [9] and in our earlier work [10] to eliminate the need for reference temperature control. In this study, we fabricated a microfluidic chip sensor by integrated a thin film thermopile sensor on to a microfluidic device fabricated using rapid prototyping technique.


Fabrication of thin film thermopile

Thin film thermopile with antimony (Sb) and bismuth (Bi) metals (Sigma Aldrich) was fabricated using thermal evaporation technique. Complementary metal shadow masks (Towne Technologies) were used to fabricate the thermopile structure. First, a Bi layer of thickness 0.8 [micro]m with a mask pattern is deposited and a complementary pattern is aligned and then a Sb layer of thickness 1.2 [micro]m is deposited on a glass coverslip of thickness 170 um (Electron microscopy sciences). Second, a metal mask with a pattern for the connector leads is also fabricated on the same coverslip by depositing 1.2 [micro]m Sb layer. Thermopiles were also fabricated on 100 um thick Kapton[R] sheet ( This fabrication procedure described in detail in ref [10]. The fabricated thermopiles typically have a resistance of 20 K[OMEGA] and showed a Seebeck coefficient of 7.14 [micro]V [(mK).sup.-1].

Microfluidic chip sensor (MCS) fabrication

A microfluidic chip sensor (MCS) is fabricated by integrating the thermopile on the glass coverslip as the channel wall of the microfluidic device. Inlet holes (Inlet 1 and Inlet 2) and an outlet hole are drilled onto the microscope glass slide along the length. The microfluidic device is fabricated using an inexpensive rapid prototyping technique called Xurography. A dual side adhesive tape ( is cut to form the channel and is sandwiched between a microscope glass slide and a glass coverslip (thermal conductivity, k-1.05 W [(m.K).sup.-1]) with the fabricated thermopile. Two configurations of the microfluidic chip sensors (MCS-1, MCS-2) were fabricated, tested and evaluated. MCS-1 was fabricated by sandwiching the dual side adhesive tape between the microscope glass slide and the glass coverslip. A thermopile sensor is attached to the coverslip using superglue, and the thermopile is facing outside the channel (Fig. 1A). Attaching a 25 um thick tape passivizes thermopile facing outside the channel. MCS-2 is fabricated by sandwiching a dual side adhesive tape with the glass coverslip with thermopiles, which is coated with a 3-um thick SU-8 (Microchem) photoresist layer and the microscope glass slide. The thermopiles in the MCS-2 configuration are facing inside the microfluidic channel and the SU-8 (thermal conductivity, k-0.2 W [(m.K).sup.- 1]) layer passivivates the thermopiles (Fig. 1B). MCS-2 fabrication was an improvement to the fabricated MCS-1.

Experimental procedure

The experimental setup used for this study consists of syringe pumps, which continuously flow water into the inlets of the MCS. An injection valve is used to inject the sample into the inlet 2 flow for generating a chemical reaction. The inlet 2 flow is hydro-dynamically focused into the MCS. The reaction is generated over the measuring junctions of the thermopile, which generates a proportional voltage. This voltage is detected by a nano-voltmeter (Agilent 34420A) and is recorded in a computer. A 13 [micro]l sample volume injected into the flow is injected using the sample injection valve. To demonstrate the operation of the MCS, the exothermic nature of glycerol water mixing reaction is utilized.


MCS-1 response to glycerol-water mixing

The fabricated MCS-1 was connected to the experimental setup and varying concentrations of glycerol were injected to measure the response of the thermopile. Flow rates of 100 [micro]l [min.sup.-1] and 25 [micro]l [min.sup.-1] were used to continuously inject the water in inlet 1 and 2 respectively. Once the sample was injected into the inlet 2 flow stream, the sample traveled to the microfluidic device and was hydrodynamically focused over the measuring junctions of the thermopile. The glycerol sample mixed with the water flowing from inlet 1 at the interface and generated heat. As the sample passed over the measuring junctions of the thermopile, the voltage increased and then returned to its baseline once the sample flowed past the thermopile. Fig. 2 shows the typical response of the thermopile for 10% (V/V) glycerol. The magnitude of the signal represents the total temperature change detected by the thermopile, and the area under the curve (AUC) represents the total heat detected by the thermopile. MCS-2 response to glycerol-water mixing

The MCS-2 device fabricated with thermopiles passivized with a 3 [micro]m layer of photoresist facing inside the microfluidic channel was also tested for glycerol-water mixing reaction. Varying concentrations of glycerol were injected into the MCS-2 and the response of the thermopile was recorded. Fig. 3 compares the response of MCS-1 and MCS-2 for 10% V/V glycerol. Flow rate dependency of the MCS-2 sensor was also investigated for flow rates of 100 [micro]l [min.sup.-1] and 25 [micro]l [min.sup.-1], and 50 [micro]l [min.sup.-1] and 25 [micro]l [min.sup.-1] inlet 1 and inlet 2 respectively (Fig. 4). The sensitivity of the sensors (in terms of AUC) MCS-1 and MCS-2 were evaluated and plotted in Fig. 5. MCS-2 showed 3.5-fold increase in sensitivity compared to MCS-1.


Both MCS-1 and MCS-2 showed responses for the exothermic reaction of glycerol-water mixing. The heat power sensitivity of the fabricated thermopile was characterized in our earlier paper as 0.045 V [W.sup.-1]. The lowest concentration of glycerol injected was 0.1% V/V, for which a voltage of 400 nV response was recorded by MCS-2. MCS-1 measured no voltage for 0.1% (V/V) glycerol concentration, because of the high thermal resistance for heat transfer to the thermopile. Considering the Seebeck coefficient of the thermopile, 400 nV corresponds to a detected temperature of 57.2 [mu][degrees]C. Similarly, considering the heat power sensitivity of the thermopile, the lowest heat power detected by the thermopile was calculated to be ~8.8 pW. MCS-2 showed better sensitivity compared to MCS-1, due to the thermal resistance in the heat transfer region for MCS-1 is higher compared to MCS-2. The total thermal resistance in the MCS-1 is due to the thermal resistance of the glass coverslip, which is 6.07 K[W.sup.-1], and the total thermal resistance in the MCS-2 only the 3 um thick SU-8 layer which is 0.833 K[W.sup.-1].


A thermoelectric microfluidic chip sensor was developed to measure dynamic temperature changes in the order of [10.sup.-6] K. High sensitivity was obtained by an inexpensive and easy to fabricate MCS without any extreme measures and complex procedures to control reference temperatures. Two configurations of the sensors were fabricated and tested MCS-1 and MCS-2. An improved sensor with low thermal resistance (MCS-2) showed a low heat power detection of 8.8 pW. The sensitivity obtained with the simple to fabricate MCS is suitable for monitoring bioprocesses such as cell metabolism, enzymatic reactions, and binding event measurements.


The authors would like to thank the Institute for Micromanufacturing at Louisiana Tech University for their support.


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Varun Kopparthy (1,2), Joshua Nimmala (1,2), Eric. J. Guilbeau (1,2)

(1) Center for Biomedical Engineering and Rehabilitation Science, (2) Institute for Micromanufacturing, Louisiana Tech University, Ruston, LA, 71272.
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Author:Kopparthy, Varun; Nimmala, Joshua; Guilbeau, Eric. J.
Publication:Journal of the Mississippi Academy of Sciences
Article Type:Report
Geographic Code:1USA
Date:Apr 1, 2014
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