MX2012014893A

MX2012014893A - System for quantifying, by dielectrophoresis and surface plasmon resonance, concentrations of particles in fluidic suspension and control method.

Info

Publication number: MX2012014893A
Application number: MX2012014893A
Authority: MX
Inventors: Hector Rafael Siller Carrillo; Jose Israel Martinez Lopez; Victor Coello Cardenas; Rodolfo Cortes Martinez
Original assignee: Itesm
Priority date: 2012-12-17
Filing date: 2012-12-17
Publication date: 2014-06-24

Abstract

The system for quantifying, by dielectrophoresis and surface plasmon resonance, concentrations of particles in fluidic suspension and control method, comprises a coupled system of detection, quantification and manipulation of particles. The configuration of this system, as well as the operation method of the same, allows the quantification and manipulation of the particles to be reviewed to be more efficient than other state-of-the-art devices, since the use of the plasmon resonance, along with insulation structures, electrodes, all within a sample cell, allow the particles to be concentrated in a predetermined area, which contributes to an efficient analysis of the same.

Description

System for quantifying, by means of dielectrophoresis and surface plasmon resonance, particle concentrations in fluidic suspension and its control method OBJECT OF THE INVENTION The system allows the detection, quantification and manipulation of inert and biological particles through the interaction of a coupled system of detection, quantification and manipulation of particles. Through the phenomenon of surface plasmon resonance, changes in the refractive index of a material, produced by the presence of inert and biological particles, are detected and quantified; these can be manipulated in the system through the dielectrophoretic force produced by a structure of insulating posts.

BACKGROUND The patent application US2004 / 0026250 Al proclaims the reduction of the number of electrodes required to create non-uniform electric fields that allow to produce dielectrophoresis using insulating structures. The invention described in this document differs in that it has the ability to measure the concentration of particles in a given space and delimit, under the effect of dielectrophoresis, by means of a sensor system.

The patent application US 2010/0224493 Al indicates the device and method for handling particles without the need for electrodes to be in direct contact with a voltage source. In contrast to the invention described in this document, the system is capable of measuring the concentration of the particles under the effect of dielectrophoresis in a determined and delimited space.

Patent JP 200534502 A points out how to measure components of a sample through the resonance of an emission source in a diffraction grating. By applying a voltage between two points, the components of the sample are attracted to the detection zone where components are detected by changes in light intensity of zero order. The invention described in this document differs by using a dielectrophoretic force generated by arrangement of insulating structures for the process of measuring particles in a given and delimited space.

Dielectrophoresis (DEF) is a non-destructive electrokinetic transport phenomenon in which the movement of particles is caused by the polarization effects of a non-uniform electric field (Pohl, 1951). The creation of non-uniform fields has been developed in two different directions: generating DEF through electrodes and with insulating structures (Cummings and Singh, 2003). So far, the first trend is the one with the most background. Through microelectrodes that generate electric fields of alternating current it is possible to take advantage of the differences of the dielectric properties to separate biological particles. For example, Morgan et al. Managed to separate the Tobacco Mosaic Virus (TMV) and the Herpes Simplex Virus (VHS) (Morgan et al., 1999) from a mixture; Gascoyne and colleagues separated cancer cells from the blood (Gascoyne, et al., 1997); Müller et al. Incorporated three-dimensional microelectrodes to separate eukaryotic cells and latex particles (Müller, et al., 1999).

To understand the basic concept of the operation of the proposed invention, the following information is required: Surface plasmon sensor (SPR for its acronym in English surface plasmon resonance). A surface wave SPP (Surface plasmon polarization) is an electromagnetic wave that propagates between the interface of a dielectric and a metal. The SPP is a transverse magnetic wave TM whose vector is parallel to the plane of the interface and is characterized by the propagation constant and the distribution of the electromagnetic field. The SPP wave propagation constant can be expressed as: Equation 1: Where ? is the angular frequency, c is the speed of light in vacuum, eD and eM are the dielectric functions. This equation describes the SPP when the real part of e? is negative and the absolute value is less than sD.

Due to the high energy losses of the metal, the SPP propagates with high attenuation in the visible and near infrared spectral regions. Since most of the field of an SPP is concentrated in the dielectric, the propagation constant of the SPP is extremely sensitive to changes in the refractive index of the dielectric material. The principle of recognition of biomolecules lies in the fact that biological receptors such as antibodies or a DNA sequence that adhere directly or through a biological matrix to the surface of the metal and capture analytes present in a liquid sample produce an increase in the rate of refraction of the metal.

The increase of the spare index gives rise to an increase in the propagation constant of SPP on the surface of the metal, which can be measured by optical and optical-electronic means as in a CCD sensor or in a spectrophotometer, among others.

The magnitude of the change in the propagation constant of the SPP depends on the change in the refractive index of the light at the metal-dielectric interface and its distribution with respect to the profile of the SPP field. There are two extreme cases: 1. The capture of the analyte occurs at a short distance from the metal surface. If the adhesion event occurs within the corresponding distance d in the real part of the propagation constant it can be expressed as follows: Equation 2 Where and ns denote the refractive index of the biolayer and the medium, respectively. The change in adhesion induced in the propagation constant of the SPP is proportional to the change in refractive index and depth. Factor F adjusts the fact that the interaction acts only on a fraction of the SPP field.

According to the theory of disturbance, the change for this case: 1. The capture of the analyte occurs at the end of the SPP. The change for this case produces a real change in the constant of proportional type. Thus: Equation 3: Re { Ap.}. = k n Where k denotes the wave number of the free space.

On the other hand, a surface plasmon sensor (SPR) comprises an optical system, a transducer means that connects a biochemical domain to the optical and an electronic system that processes the sensor information.

Another process used in the system is Dielectrophoresis by insulating posts (IDEP), where the dielectrophoretic force acting on a spherical particle can be described by the following equation: Equation 4.

FDEF = 2nE0emr3Re [f (api am)] V (£ | E) where : e0 is the permittivity of free space, em is the relative permittivity of the suspension medium, r is the radius of the particle, (£ || E) is the intensity of the electric field and f [pv, it is the Clausius-Mosotti factor (CM), given in equation 5: Equation 5: describes the electrostatic properties of a particle and where: < ? p and dm are the complex conductivities of the particle and the medium respectively, which are defined as described in the following equation: Equation 6 s = s + '? e Where: ? is the angular frequency of the application of the electric field, (Jones, 1995).

As observed in Equations (l) - (3), the forgetic dielectric force exerted on a particle depends on the electric field strength, the particle size, the dielectric properties of the particle, as well as the conductivity of the particle. suspension medium. These operating conditions can be manipulated to increase / decrease the dielectrophoretic force exerted on a particle, and thereby achieve to separate and / or concentrate a specific type of particle. Due to this flexibility in operating conditions, dielectrophoresis represents an excellent option for the concentration and manipulation of particles (Lapizco-Encinas, et al., 2005).

In many systems, when operating with low frequency AC electric fields (less than 100 kHz) or direct current fields, it is considered that the Clausius-Mossotti factor can be calculated using the actual conductivities of the particles and the medium ( Hughes et al., 2001).

The conductivity of the particle can be calculated with the following equation: Equation 7 2Ks s? = ° b + - Where: ab is the conductivity of the material of the particle, Ks is the conductance of the surface and r is the radius of the particle.

The dielectrophoretic force acting on a particle can be positive or negative according to the sign of the Clausius-Mossotti factor. If the polarization of the particle is greater than that of the immersion medium, the induced dipole is parallel to the electric field and the particle is attracted by regions of greater field strength. Under these conditions the particles adhere to the edges of the electrodes or concentrate in the narrow part of the insulating posts. On the other hand, if the polarization of the particle is smaller than that of the immersion medium, the induced dipole is antiparallel to the electric field and the particle is repelled by regions of intense field. The particles are concentrated in the center of the electrode arrangement or in the center of the insulating posts (Gascoyne and Vykoulal, 2002). Unlike electrophoresis, dielectrophoresis has a second order dependence with respect to the electric field E, so at low values of electric field the magnitude of the dielectrophoretic force will be negligible. If there are similar values between the electrokinetic and dielectrophoretic forces when increasing the field, there will be a movement of bioparticles known as dielectrophoresis of currents, increasing the value that will cause the dielectrophoretic force to overcome the electrokinetic, diffusion, Brownian movement and we will find the trapping dielectrophoresis regime where the particles are concentrated and immobilized (Cummings and Sing, 2003), as in equation 7: The dielectrophoretic mobility equation ^ DEF) defines the factors that determine the presence of a flow with dielectrophoretic regimen or that of trapping dielectrophoresis by means of equation 8: r2 £ mRe [f (ap, a)] 3? Where r is the radius of the particle, emes the relative permittivity of the medium and 7 is the viscosity of the medium.

BRIEF DESCRIPTION OF THE FIGURES Figure 1. Isometric view of the system to quantify concentrations of particles in fluid suspension by means of dielectrophoresis and surface plasmon resonance. Figure 2. Schematic representation of the operation diagram of the detection and manipulation system.

Figure 3. Diagram of geometries of the arrangement of insulating structures.

DETAILED DESCRIPTION OF THE INVENTION The invention comprises an integrated system for detecting and manipulating particles using surface plasmon, as well as dielectrophoretic forces caused by non-uniform electric fields, mentioned above, in the background section.

The system first proposes a device for the detection and manipulation of particles (see figure 1), which comprises: a sample cell (12) either of glass material, polymer or composed of glass and polymer or by the combination of both, which presents an entrance port (7) connected through a hose or tube, to a tank (10), where a liquid sample is placed and with the help of at least one system pumping (2) consisting of at least two piezoelectric or mechanical pump controllers, is carried to the inlet port (7) of the sample cell (12); the sample cell (12) functions as a channel, through which the liquid sample transits from one end to the other at a certain speed; the system also comprises at least one microcontroller (8) that stores a control program to activate the emission of a coherent single-phase light, such as a solid-state diode or a laser (1) that reaches a detection zone (3) of gold or silver or some other material, deposited with elements of biological recognition, and where the detection zone (3), is inside the sample cell (12) on the surface opposite an arrangement of insulating structures (9), which, is at the mediation of the sample cell (12) that is, mediation of the channel. The arrangement of insulating structures (9), is made of dielectric material such as polymethylmethacrylate (PMMA), polydimethylsiloxane (PDMS) or SU8, and where said insulator structures (9) have a three-dimensional geometry with curved elements either poles or half spheres with right angles as step barriers or zigzag structures (see Figure 3). In addition, the arrangement of insulating structures (9) occupies a partial or total fraction of the height of the channel, which favors the generation of non-uniform electric fields by the variation of the electric field value along the channel, and the concentration of particles in the detection zones, by increasing the active dielectrophoretic force in specific regions of the sample cell (12). In the sample cell (12), a contact surface is located of dielectric material, either glass or some polymer, where said contact surface is located below the conductive region (6), and generates a variant optical signal to the angle of incidence of the light and proportional to the amount of biological elements of entrapment occupied on the detection zone (3), and that is received and converted to an electrical signal by an optoelectronic transducer (4), for example, a CCD camera or a diode array, and stored in the microcontroller (8).

The microcontroller (8) also has the task of measuring the refractive angle that can be associated with the value of the concentration of the particle of interest, varying the angle condition of the sample excitation light with respect to the light source through of a high-precision stepper motor (13) rotating the sample cell (12) on its own axis, or modulating the excitation light (1) in phase by varying the intensity of the light.

The sample cell (12), has an exit port (1 1), at the opposite end to the entrance port (7), where the sample can be purged to allow sampling continuously, as well as for operations cleaning the cell itself. A substrate or electrode (5) of different polarity and preferably of the same dielectric material as the insulator structures (9), is present inside the sample cell (12) at each of the ends of its sides of shorter length, is say, near the ports of entry and exit respectively, and where said substrate is intended to supply the energy to increase the magnitude of the electric field and cause the dielectrophoretic force in the areas adjacent to the region of insulating structures (9) to be increase and favor the accumulation of particles in the detection areas of biological recognition.

The control method of the system for quantifying, by dielectrophoresis and surface plasmon resonance, particle concentrations in fluid suspension consists of the following steps: a) Place the liquid sample in the tank (10); the tank is connected to the inlet port by a hose or tube; b) Start flow of the sample; the microcontroller (8) initiates the flow of the sample towards the interior of the sample cell (12) through the activation of a piezoelectric or mechanical pump, where said sample has a flow velocity of 5 to 50μ1 / p ??; c) Activate and vary the conditions of incidence of the excitation light; the microcontroller (8) varies the incident conditions of the excitation light, through a controlled rotation of the sample cell (12) or of the phase modulation of the intensity of the excitation light; the variations are recorded in an optoelectronic system (CCD camera, diode array, etc.) based on a preset program by the user. d) Supply voltage to the electrodes; the microcontroller (8) supplies the direct current voltage to the electrodes (5) simultaneously, in the range of 50 to 500 V to increase the electric field and the dielectrophoretic force, and promote the transfer of sample to the detection zone (3). e) Detect and quantify the particles of interest; under the influence of the pumping system (2) and the interaction of the electric field with the insulating structures (9) that produce the dielectrophoretic force, the particles of interest contained in the liquid sample, adhere to the biological surface, found in the area detection (3) causing a change in the refractive index recorded by an optoelectronic system. f) Record the changes in the refractive index of the metal-dielectric interface; these changes are recorded through the optoelectronic system (CCD camera, diode array, etc.) in the microcontroller (8) where they are associated with concentration values for the particle of interest. g) Save and send information; the microcontroller (8) is responsible for storing the information and sending it to a user interface (14) which may consist of a personal computer or a screen. h) Finish the pumping cycle; the microcontroller (8) orders the completion of the pumping cycle and turns off the light excitation signal and the voltage signal of the electrodes (5), whereby the sample is purged until the sample cell is emptied.

Claims

CLAIMS Having described my invention enough, I consider it as a novelty and therefore claim as my exclusive property, what is contained in the following clauses:

1. A system for quantifying by means of dielectro foresis and surface plasmon resonance particles of biological interest in fluidic suspension characterized in that it comprises a sample cell, connected to a tank, where said sample cell has an inlet port and an outlet port, arranged opposite way, so that a fluidic sample transits through it, and where in each of the ports, an electrode of opposite polarity is located; between the input and output ports, a detection zone and an array of insulating structures are aligned; in the port of entry to the sample cell a pump system is connected, by means of which the sample is flowed through the body of the cell, at a determined speed; said pumping system is controlled by a microcontroller. which in turn is connected to an interface located below the conductive region for general an optical signal variant to the angle of incidence of light.

2. The system for quantifying by means of dielectrophoresis and surface plasmon resonance particles of biological interest in fluid suspension according to claim 1, characterized in that the sample deposit is aligned with the inlet port of the sample cell through a hose or tube .

3. The system for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension according to claim 1, characterized in that the microcontroller stores a control method to activate the emission of a coherent single-phase light on the sample, said light comes from a light source, preferably laser or a laser diode.

4. The system for quantifying by means of dielectrophoresis and surface plasmon resonance particles of biological interest in fluid suspension according to claim 1, characterized in that the detection zone is optionally gold or silver.

5. The system for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension according to claim 1, characterized in that the insulating structures have a three-dimensional geometry with curved elements, which can be poles, half spheres with right angles as barriers of step or zigzag structures.

6. The system for quantifying by means of dielectrophoresis and surface plasmon resonance particles of biological interest in fluid suspension according to claim 1, characterized in that the output port of the sample cell allows continuous sampling of particles, that the cell be purged, as well as as the operation of cleaning it.

A control method of the system for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension in accordance with that described in claims 1 to 6, characterized in that it comprises the following steps: a) Place the liquid sample in a tank; b) Start flow of the sample; c) Activate and vary the incidence conditions of the excitation light; d) Supply voltage to the electrodes; e) Detect and quantify the particles of interest; f) Record the changes in the refractive index of the metal-dielectric interface; ' g) Save and send information; h) Finish the pumping cycle;

The control method of the system for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension according to claims 1 to 6, characterized in that in step b) the microcontroller initiates the flow of the sample into the interior of the sample cell through the activation of a piezoelectric or mechanical pump, where said sample has a flow velocity of 5 to 50μ1 / p ??

9. The control method of the system for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension according to claims 1 to 6, characterized in that in step c) the microcontroller varies the conditions of incidence of the excitation light, through a controlled rotation of the sample cell or in-phase modulation of the intensity of the excitation light;

10. The control method of the system for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension according to claim 9, characterized in that in step c), the variations are recorded in an optoelectronic system (CCD camera, diodes, etc.) based on a program preset by the user.

1 1. The system control method for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension according to claims 1-6, characterized in that in step d) the microcontroller supplies the direct current voltage to the electrodes simultaneously, in the range of 50 to 500 v to increase the electric field and the dielectrophoretic force, and encourage the transfer of sample to the detection zone.

12. The control method of the system for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension according to claims 1-6 characterized in that in the stage e), the pumping system with the interaction of the electric field with the insulating structures that produce the dielectrophoretic force, the particles of interest contained in the liquid sample, adhere to the biological surface, found in the detection zone, causing a change in the refractive index that an optoelectronic system registers.

13. The control method of the system for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension according to claims 1-6, characterized in that in step f) the changes are recorded through the optoelectronic system in the microcontroller, where they are associated with concentration values for the particle of interest.

14. The control method of the system for quantifying, by means of dielectrophoresis and surface plasmon resonance, particles of biological interest in fluid suspension according to claims 1-6, characterized in that in step g), the microcontroller is in charge of storing the information and sending it to a user interface that can consist of a personal computer or a screen.

15. The method of control of the system for quantifying by means of dielectrophoresis and surface plasmon resonance particles of biological interest in fluid suspension according to claims 1-6, characterized in that in step h), the microcontroller orders the completion of the pumping cycle and turns off the signal of light excitation and the voltage of the electrodes, with which, the shows until the sample cell is empty.