JP2933362B2 - An improved system for measuring electrokinetic properties and characterizing electrokinetic separation by monitoring current in electrophoresis - Google Patents

Description

The present invention relates generally to a capillary electrokinetic device, and more particularly to the measurement of electrokinetic properties and the characterization of electrokinetic separation by monitoring current during electrophoresis. An improved system for performing.

BACKGROUND OF THE INVENTION In capillary zone electrophoresis (CZE), an ionic transfer channel of capillary size is filled with an electrolyte.
The sample to be analyzed is injected at one time in the channel, and a high voltage is applied across the channel, so that the sample inside the electrolyte moves from one end of the channel to the other.
If the electrolyte is in contact with the capillary wall, the inner surface of the capillary is charged through ionization of the surface groups on the capillary wall or through the absorption of charge particles from the electrolyte onto the inner surface. In each case, the electrolyte inside the capillary is not electronically neutral,
Acquire a positive or negative net charge. Under the action of the applied electrolysis, the electrolyte moves towards one end of the capillary. This movement is called electroosmotic flow.

While the electrolyte flows generally towards one end of the capillary and the electrolyte acquires a net charge as a whole,
Each component of the sample is positively or negatively charged, respectively.

Therefore, in addition to the electroosmotic flow of electrolyte, electrophoresis also occurs. That is, the applied electric field applies a force for moving the positively charged particles toward the negatively charged electrode and moving the negatively charged particles toward the positively charged electrode. As a result, the components in the injected sample separate into zones that are classified based on their mobility.

Often, the speed of the electrophoretic flow is less than the electroosmotic rate. As a result, the particles in the injected sample move in one direction, the direction of the electroosmotic flow, and as each band passes through a suitable detector located below the capillary inlet, Various particles can be detected.

Obviously, accurate characterization of the electroosmotic flow is strongly desired not only for understanding CZE, but also for optimizing the action of CZE in analyzing a given sample. One way to measure electroosmotic velocity is to record the transit time of the injected uncharged marker solute. This uncharged marker solute is carried through the capillary under the action of only electroosmotic flow. This technique is described by Warburol and Jojienson in "Analytical Chemistry, Vol. 58, pp. 479-481, published in 1986." Capillary band electrophoresis "(see reference line 1, and reference 2).

First, it is extremely difficult to select a truly neutral marker. Further, the charge on the particular marker selected depends on the medium on which it is located. Therefore, an apparently "neutral" marker used in one medium is not suitable for use as a marker in another medium. In addition, it is difficult to detect "neutral" markers.

Another method of measuring the electroosmotic rate consists in measuring the weight of the electrolyte transported from the capillary inlet to the capillary outlet over the timed time interval. For a description of this method, see Altair and Simpson, Vol. 23, pp. 453-454 of the Analytical Process Journal (ANAL. PROC), published in 1986, entitled “High Voltage Capillary Band Electrophoresis. Measurement of Electroosmotic Flow ". In this method, the loss due to evaporation must be eliminated by coating the liquid surface in the cathode compartment with a thin film of silicon fluid or by using an airtight lid. The weight of the electrolyte transported is so small that it seems necessary to use digital equilibrium. Measuring a light weight transport is inconvenient and cumbersome, and special care must be taken to ensure the accuracy of this measurement. However, this method requires that a small amount of transport be measured.

In addition, the inventors of the present invention have published an analytical chemistry journal published in 1988,
Volume 60, pp. 1837-1838, written by Fang, Gordon, and Zear, on a basic method entitled "Current Monitoring Method for Measuring Electroosmotic Velocity in Capillary Band Electrophoresis". Has been announced.

None of the above methods are satisfactory in forming a system. It is therefore desirable to provide an improved system for measuring the electrokinetic properties of a medium, which alleviates the above difficulties.

(Means for Solving the Problems) The apparatus of the present invention is for determining the electrokinetic properties of the first electrolyte in the electrokinetic separation. The device comprises a long capillary having an inlet end and an outlet end suitable for holding an electrolyte, a second electrolyte suitable for introduction into the capillary from an inlet end after the first electrolyte has been introduced. Consists of The second electrolyte has a specific resistance different from that of the first electrolyte, but has substantially the same chemical properties as the first electrolyte. A voltage is applied to the electrolyte and a current is applied to the electrolytic chamber in order to cause an electroosmotic flow of the electrolyte in the direction of the outlet end and to continuously replace the first electrolyte in the tube with the second electrolyte. Means for flowing are also provided in the device. The device further comprises means for measuring the current passing through the electrolyte in the capillary as a function of time. The electroosmotic rate of the first electrolyte is determined by measuring the current as a function of time.

In one embodiment, the first electrolyte (Electrolyte 1) comprises a first solution comprising a buffer, and the second electrolyte (Electrolyte 2) comprises the same buffer having a concentration different from that of the second solution. Consists of a liquid. The capillary is initially filled with electrolyte 1. Next, the electrolyte 2 is continuously
The electrolyte 2 is continuously injected so that the electrolyte 2 is replaced by the electrolyte. By measuring the change in current across the capillary as a function of time, the rate of electroosmosis is determined. In this process, it is assumed that electrolyte 1 and electrolyte 2 have slightly different resistances but basically have similar electroosmotic rates.

Once the electroosmotic rate is established, the electroosmotic transfer rates of the multiple components in the sample are determined as follows. First, the capillary is filled with electrolyte 1. Next, a thin strip of sample is injected into the capillary from the inlet. Therefore, electrolyte 1
Is injected electroosmotically. It is assumed that the presence of multiple components in the sample solution does not significantly alter the electroosmotic rate of electrolyte 1. By using the knowledge of the electroosmotic velocity of the electrolyte 1 and by measuring the transit time during which multiple components in the sample solution are transported from the capillary inlet to the detector area of the capillary, the electroosmotic velocity of the multiple components is determined. Can be determined. In addition, using the knowledge of the interface strength inside the capillary (usually given by the ratio of the voltage applied across the tube to the length of the tube), the electrophoretic mobilities of the various components can be derived from the electrophoretic velocities. You.

The second embodiment is related to the first embodiment, but the electroosmotic speed of the electrolyte and the electrophoretic speed of a plurality of components in the sample solution are measured in the same experiment. Is done.

This is performed as follows. First, the capillary is filled with electrolyte 1. Next, a thin strip of the sample solution is injected into the capillary inlet.

Next, the electrolyte 2 is electrokinetically injected into the inlet end of the capillary so that the electrolyte 1 is replaced by the electrolyte 2. In addition to measuring the transit time during which each component in the sample solution is transported a known distance from the entrance end to the detection area of the capillary, the change in current across the capillary as a function of time is measured to determine the individual sample components. In addition to determining the rate of electrophoresis, it is possible to determine the rate of electroosmosis of the electrolyte. The electrophoresis speed of a plurality of components can be determined from the electric field strength in the tube, and the electrophoresis speed can be determined in the same manner as in the first embodiment.

(Embodiment) FIG. 1 is a system diagram of a system for determining an electroosmotic speed for explaining an embodiment of the present invention. Within the capillary tube 14, the rate of electro-osmosis of the first electrolyte is determined, and as shown in FIG.
It is immersed in. The other end of the pipe 14 is immersed in the second storage tank 16. For the purpose of measuring the electroosmotic rate of the capillary 14 and the first electrolyte inside the storage tank 12, the electrolyte inside the storage tank 16 has virtually the same chemistry as the electrolyte inside the storage tank 12. However, the specific resistance is different from the specific resistance of the electrolyte inside the storage tank 12. Voltage source
A high voltage is applied between the two storage tanks from 18, which places the second electrolyte inside the storage tank 16 into the capillary tube 14,
The first electrolyte existing inside the tube 14 is continuously replaced with the second electrolyte until all of the first electrolyte inside the tube 14 is replaced with the second electrolyte.

In order to measure the change in the current flowing through the electrolyte in the tube,
A resistor 20 is connected in series with the two reservoirs. Therefore, the change in current through the two electrolytes inside the two reservoirs and tubes 14 is recorded by the chart recorder 22 as a voltage change across the resistor 20. The second electrolyte inside the storage tank 16 enters the tube 14 and the first electrolyte originally inside the tube 14
When a portion of the electrolyte is replaced by a second electrolyte, the resistance of the electrolyte column inside the tube 14 varies in resistivity because the specific resistances of the two electrolytes are different. The magnitude of the current in the circuit through the two electrolytes thereby changes.

For example, if the second electrolyte has a higher resistivity than the first electrolyte being replaced, after some of the first electrolyte is replaced by the second electrolyte inside the tube 14, The resistance of the electrolyte column inside tube 14 increases. Therefore, the current through resistor 20 decreases and the voltage recorded by recorder 22 drops by a corresponding amount. For example, this is shown in the graph of FIG.

That is, even if the electrolyte inside the storage tank 16 is small,
Prior to entry, the first electrolyte inside the storage tank 12
If is satisfied, the current starts to flow. Recorder
The current recorded by 22 is the current shown in the graph of FIG.

Thus, the current through the resistor 20 decreases until all of the first electrolyte inside the tube 14 is replaced by the second electrolyte inside the reservoir 16. At this point, the resistance of the electrolyte column inside tube 14 does not increase so that the current recorded over all time is kept constant.

From the above, the start time of recording time t ₀ coincides with the point in time at which the second electrolyte enters tube 14 to replace the first electrolyte with the second electrolyte, and time t ₁ is the time at which the second electrolyte enters tube 14. This coincides with the point in time when all of the one electrolyte has been replaced by the second electrolyte inside the reservoir 16. Therefore, the time difference between t ₀ and t ₁ is the time required for all of the first electrolyte inside tube 14 to be replaced by the second electrolyte. In other words,
During the period (t ₁ −t ₀ ), the first electrolyte flows in the length direction of the capillary 14. Since the length of the capillary 14 can be easily measured, the electroosmotic rate of the first electrolyte is thereby reduced by the tube 14.
Is determined by the ratio of length to time (t ₁ -t ₀ ). The time interval (t ₁ −t ₀ ) also fills the capillary with the second electrolyte, and the second electrolyte completely replaces the first electrolyte, so that the second electrolyte is inserted into the capillary. Is a period during which flows. Therefore, the electroosmotic rate of the second electrolyte is also determined by the ratio of the length of the tube 14 to the time (t ₁ -t ₀ ). In effect, two or more electrolytes having the same chemistry are continuously introduced into tube 14 in the same manner as measuring the electroosmotic rate of multiple electrolytes. In such a case, the plot of time versus current obtained from such a measurement represents a more changing point (point where the slope of the graph changes) than the plots of FIGS. 2 (A) and 2 (B).

Because the two electrolytes have the same chemistry,
These electrolytes effectively have the same effect on the inner surface of the capillary. This eliminates what is likely to be one of the likely sources of error when measuring the electroosmotic rate. In this embodiment, the second electrolyte is the same as the first electrolyte, but has a different concentration. In one study considered to be satisfactory, the first electrolyte consists of a 20 mM sodium phosphate buffer with a pH of approximately 7.0. The second electrolyte is prepared by diluting the first electrolyte with water such that the ratio of electrolyte to water is 19: 1 so that the concentration of the second electrolyte is 95% of the concentration of the first electrolyte. Generated.

Thus, the concentrations of the two electrolytes need to differ by only a small amount to cause a measurable difference in the current sensed through resistor 20. It is also conceivable that the second electrolyte is different from the composition of the first electrolyte. All such modifications are also included in the scope of the present invention.

In the above embodiment, the two electrolytes differ only in the concentration of the dissolved substance. The question that arises is whether electroosmotic rate is a strong function of electrolyte concentration. For this purpose, the reservoir 16 is filled with 20 mM sodium phosphate buffer, and in addition to the tube 14, the reservoir 12 is instead filled with 19 mM sodium phosphate buffer. Again, the same voltage is applied from the power source 18 to both ends of the two storage tanks. The resulting plot of time versus current is shown in FIG. 2 (B). As shown in FIG. 2 (B), (t ₃ −t ₂ ) indicates that the first electrolyte in the storage tank 16 is
This is the time required to completely replace the second electrolyte inside 14. (T ₃ −t ₂ ) corresponds to (t ₁ −
t ₀ ) has been confirmed to be almost the same. Thus, electroosmotic rate is not a strong function of electrolyte concentration.

The above system, and the method for measuring the electroosmotic velocity of the first electrolyte described with reference to FIGS. 1 and 2 (A), uses the electrophoretic velocity of the sample, ie, the characteristic characterizing the sample. It is used to derive. Thus, if a sample having many components is introduced into the interior of tube 14, these components will acquire a positive or negative charge.

Therefore, the electric field applied by the power supply 18 moves the positively charged component to the negatively charged electrode,
A force is applied to the negatively charged component so as to apply a force to the positively charged component and move the negatively charged component to the positively charged electrode. Therefore, the actual velocity of each of these components is the vector sum of the two velocities.
The overall velocity of the first electrolyte, as well as the velocity of the electrophoretic flow of such components, is these two velocities.

Thus, if V is the actual velocity of the component, V _eo is the total electroosmotic velocity of the electrolyte, and V _e is the electrophoretic velocity of the component, the actual velocity V is given by V = V _e + V _eo , The electrophoretic migration speed U _e is given by U _e = V _e / E. Where E is the electric field provided by the power source 18. Therefore, if the actual speed and the electroosmotic speed of the component are measured, the applied field strength E is known, and the electrophoretic mobility of the component can be derived from the above equation.

In a conventional manner, the actual velocity of the component can be measured by employing a detector 24 located near the tube 14. Thus, if the length of the capillary section through which the component inside the first electrolyte flows to reach the detector 24 is L, the actual velocity of the component is given by V = L / Tm . Here, the sample is injected into the inlet terminal of the tube 14 until the time when the detector 24 detects the component, and Tm is this time.

Obviously, the electrophoretic mobilities of many components inside the sample can be measured in a single measurement in which one or more detectors 24, resistors 20, and chart recorder 22 are all used in a single operation. , Can be measured in a similar manner. At the entry end, if the sample is injected into the first electrolyte inside the tube 14 and, together with the sample, the first electrolyte is replaced by a second electrolyte in a manner similar to that described above, First
The electroosmotic rate of the electrolyte and the actual rate of the component in the sample are determined in a single measurement step. This is the method of implementation described above in the summary of the present invention (means for solving the problem).

Of course, it is possible to measure the moving speed of the sample components as described above in the summary of the present invention (means for solving the problem) in an independent experiment. Thus, after determining the electroosmotic rate of the first electrolyte as described above, the same capillary is filled with the first electrolyte.

The sample in which the first electrolyte is continuously present in a large amount is
Injected. In effect, the same voltage that was applied during the electroosmotic rate measurement is again applied across the tube. In order to determine the moving speed of a plurality of components over the entire known distance, the moving time of a sample component is detected.

As known to those skilled in the art, micelles can be added to the electrolyte to enhance the electrokinetic separation function. Thus, each component in the sample has a different affinity for the micelle. Therefore, the presence of micelles affects the speed of movement of such multiple components. However, in order to accurately measure the moving speed of these components, it is necessary to measure the speed of micelles as a reference.

As is known to those skilled in the art, micelles move very slowly, the speed of which is difficult to detect. The system proposed by the applicant provides a convenient way to easily determine the speed of micelle movement. This has been explained with reference to FIGS. 1 and 3.

Referring to FIG. 1, storage tank 12 and tube 14 are filled with a first electrolyte, and storage tank 16 is filled with a second electrolyte mixed with micelles. In this description, the specific resistance of the second electrolyte is higher than that of the first electrolyte so that the current increases when the first electrolyte inside the tube 14 is replaced by the second electrolyte and micelles. Low. It is understood that the micelles used are, for example, sodium dodecyl sulfate, and that the migration speed of the other micelles can be measured in a similar manner.

The increase in current shown in FIG. 3 is explained by the presence of both sodium and dodecyl sulfate ions. Sodium ions inside the second electrolyte move relatively quickly, causing an increase in current. As shown in FIG. 3, the current change rate changes at t _5, and reaches the sodium ions by time t ₅ to the other terminal of the tube 14. After time t _5, the current carried by the movement of sodium ions has become constant, the current increase are those carried by dodecyl sulphate ions.

Meanwhile, the dodecyl sulfate ions very slowly moves along the tube 14, these ions are continuous between times t ₅ and time t _6, results in a small current increment.

At time t _6, the current detected by the recorder 22 becomes constant, finally dodecyl sulfate ions by time t ₆ has reached the other terminal of the tube 14. Therefore, the time interval (t ₆ −t ₄ ) is the movement time required for the micelle ions to move in the length direction of the capillary 14. The actual velocities of the components inside the sample that are present inside the electrolyte are again the vector sum of the electrophoretic migration speed and the micelle migration speed.
The actual travel speed of the multiple components in the presence of micelles is detected using detector 24 as before. Referring to FIG. 1 and FIG. 3, the moving speed of the micelle is detected as described above. Therefore, the electrophoretic migration speed of a plurality of components is determined.

In the method described above with reference to FIGS. 1 and 2 (A), the various currents measured at each time are recorded on a plot of current versus time as data points, and the current is plotted as a function of time. A line segment to represent is drawn. Where a data point deviates from the drawn line segment, the difference between the current value indicated by the graph drawn as the data point is the error term for such data point. Such error terms are minimized by conventional methods. In one conventional method, for example, a graph is drawn so that the above error term is minimized. As is evident from FIGS. 2A and 2B, the data points at which the current is constant are not always clearly defined. To improve the accuracy of the measurement, data points recorded at or near the point where the current becomes constant are discarded, and the graph is redrawn so that the error term is minimized until the total error is minimized. I do.

In the method described above with reference to FIGS. 1 and 2A, to measure the time required for the first electrolyte to travel the entire length of the tube 14, the first One must wait until all of the electrolyte has been replaced by the second electrolyte.

Often, this takes considerable time. The inventor proposes an alternative as described below.

The reservoir 10 and the tube 14 are filled with a first electrolyte, and the reservoir 16 is filled with a second electrolyte. Power supply 18 applies a voltage between the two reservoirs to replace the first electrolyte inside tube 14 with the second electrolyte inside reservoir 16. The current becomes constant over time, and the first
Instead of waiting until it indicates that all of the electrolyte has been replaced by the second electrolyte, the current is measured only for a short period of time. Before or after the above process, the tube 14 is filled with a second electrolyte and a power supply 18 is used to apply the same voltage between the two ends of the tube 14, causing the electrolyte to flow in the direction of one end of the tube. ing. However, the replaced second electrolyte is replaced by the same second electrolyte so that the measured current is constant. Together, these two measurements can determine the time required for the first electrolyte to travel along the length of the tube 14, as described below.

For example, tube 14 and reservoirs 12, 16 are all filled with a second electrolyte and the constant current passing through resistor 20 is I ₂
It is. There, the reservoir 12 and the tube 14 are filled with a first electrolyte, the reservoir 16 contains a second electrolyte, and the same voltage is applied between the two reservoirs by the power supply 18 as before. ing. The current values I ₁ and I ₃ are measured at times t ₇ and t ₈ , respectively. The time t ₇ simply for convenience, power supply 18
A voltage can be applied between the two reservoirs to provide a starting point for starting to replace the first electrolyte inside the tube 14 with the second electrolyte inside the reservoir 16. As shown in FIG. 4, t can be plotted so that a straight line graph 50 can be plotted.
₇ samples the current flowing through the resistor 20 in a short period of time to t ₈ from. Assuming that the rate of current change is constant throughout the process in which the first electrolyte inside the tube 14 is replaced by the second electrolyte, the point at which all of the first electrolyte is replaced by the second electrolyte, ie, the value until the current value falls into I _2, the graph extends simply as a straight line.

Therefore, t ₉ , the time when this phenomenon occurs, is
Is the time during which all the first electrolyte has moved along the length of the first electrolyte. Therefore, the ratio of the length of tube 14 to the period of (t ₉ -t ₇ ) is the electroosmotic rate of the first electrolyte.

First period (t ₇ ~t ₈₎ is for convenience, as long as that is to choose a sampling period, is understood that other sampling periods are selected in the same manner.

All such modifications also fall within the scope of the present invention. Respect graph 50, further giving more data points, in order to improve the measurement accuracy, to carry out two or more current measurements preferred between t ₇ and t _8.

Storage tank 12 and tube 14 contain a second electrolyte and storage tank 16
If contains the first electrolyte, the measurement will be as shown in FIG.

Current samples the period of the first period (t ₁₀ ~t _11),
A straight line graph 60 is plotted as shown in FIG. The current through the electrolyte is also characteristic when the tube 14 is filled completely by the first electrolyte, the value is I _1. Therefore, the total time required for the first electrolyte to enter the entire length of tube 14 and be replaced by the second electrolyte (t ₁₂
Tt ₁₀ ), so that the graph 60 can be extended again until the graph 60 crosses a straight line graph representing a current equal to I ₁ . Therefore, the electroosmotic velocity of the first electrolyte is given by the ratio of the tube L the length of the relative (t ₁₂ ~t _10).

Again, it is possible to use a different sampling period instead of (t ₁₀ ~t _11).

The above implementation has the advantage that the electroosmotic rate can be determined in a much shorter time. The current is
(T ₇ ~t ₈₎ period, or it is necessary to sample only over a short period of time such as a period of (t ₁₀ ~t _11). The constant current values (I in FIG. 4 and I 5 in FIG. ₅ ) are simple and
It can be measured quickly.

The invention has been described with reference to specific implementations and embodiments. It is understood that various modifications can be made without departing from the scope of the claims of the present invention.

[Brief description of the drawings]

FIG. 1 is a system diagram of an electrokinetic system for measuring an electroosmotic speed for explaining a first embodiment of the present invention. 2 (A) and 2 (B) are electropherograms showing the measurement of the electroosmotic velocity for explaining the embodiment of the present invention. FIG. 3 is an electrophoretogram showing the measurement of the electroosmotic velocity of an electrolyte containing micelles for explaining the present invention. 4 and 5 are electropherograms showing a preferred method for determining the rate of electroosmosis. 10 ... System 14 ... Capillary tube 12,16 ... Storage tank 18 ... Power source 20 ... Resistor 22 ... Chart recorder 24 ... Detector 50,60 ... Graph

────────────────────────────────────────────────── 72 Continuing the front page (72) Inventor Xiaofa Hang 937, Jackson Street, Mountain View, 94043, United States of America 937 (72) Inventor Jack I. Ormes United States, 94303 Aspens Street, Palo Alto, California 94877 (56) References JP-A-56-129848 (JP, A) (58) Fields studied (Int. Cl. ⁶ , DB name) G01N 27/447

Claims (9)

(57) [Claims] 1. A method for determining the electrokinetic properties of an electrolyte in an electrokinetic separation by means of a long capillary with an inlet terminal and an outlet terminal, said method comprising different electrical resistivity but substantially the same Introducing a plurality of electrolytes having chemical properties into the capillary from the inlet terminal at a time, one electrolyte at a time, and causing electro-osmosis of the electrolyte toward the outlet terminal to continuously replace the electrolyte Applying a voltage to the electrolyte to flow a current through the electrolyte; measuring the current flowing through the electrolyte as a function of time; determining at least one electroosmotic flow of the plurality of electrolytes; Performing a process of determining the rate of movement of a component of a sample in one or more electrolytes; and calculating the electroosmotic mobility of the component. 2. The step of determining a moving speed comprises steps to be performed independently of the remaining steps of the method, and introducing a first one of said plurality of electrolytes into an inlet terminal of a capillary. Injecting a sample into the capillary at the inlet end; and inserting the first electrolyte into the capillary at the inlet end even after the sample is injected to replace the first electrolyte with a sample previously introduced into the capillary. Further introducing an electrolyte, causing electro-osmosis of the electrolyte toward the outlet end, replacing the sample and the first introduced first electrolyte successively with the later introduced first electrolyte; Applying a voltage to the electrolyte to cause a current to flow through the electrolyte to move the component of the sample toward the exit terminal; andmeasuring a moving speed of the component of the sample. The method described. 3. The method of claim 2, wherein measuring the speed of movement of the component comprises measuring a time period required for the component to travel a known distance. 4. The step of determining the speed of movement comprises steps to be performed together with the remaining steps of the method, and after the first of the plurality of electrolytes has been introduced and Injecting a sample into the inlet end of the capillary before the second of the electrolytes is introduced, and wherein the applied voltage is such that, in addition to the first electrolyte, the sample also continuously flows into the second electrolyte. And moving the component of the sample toward the exit terminal, and wherein the step of determining the moving speed comprises measuring the moving speed of the component of the sample. . 5. The method of claim 4, wherein measuring the speed of movement of the component comprises measuring a time period required for the component to travel a known distance. 6. A method for determining the electrokinetic properties of an electrolyte in an electrokinetic separation by means of a long capillary having an inlet terminal and an outlet terminal, said method having different electrical specific resistances but having substantially the same chemical resistivity. A plurality of electrolytes having specific properties, one at a time, in a temporary manner such that the first of the plurality of electrolytes is introduced before the second of the plurality of electrolytes before and after is introduced. Applying a voltage to the electrolyte to cause electro-osmosis of the electrolyte toward the outlet and to continuously replace at least one of the plurality of electrolytes with another electrolyte; Flowing a current through the electrolyte to remove the electrolyte for a period of time less than the time required to completely replace the first of the plurality of electrolytes with the second of a plurality of electrolytes inside the capillary. Flow Measuring the current flowing as a function of time; filling the capillary with a second electrolyte; applying the voltage across the capillary; measuring the current flowing through the second electrolyte; Determining the electroosmotic rate of at least one of the electrolytes of the above. 7. A sample comprising one or more components, and a long capillary suitable for holding an electrolyte and having an inlet terminal and an outlet terminal; A plurality of electrolytes suitable for being introduced into the capillary from the inlet terminal in two proportions, causing electroosmosis of the electrolyte toward the outlet terminal, wherein at least one of the plurality of electrolytes is continuously transferred to another electrolyte. Means for applying a voltage to the electrolyte to cause a current to flow through the electrolyte after the electrolyte has been introduced into the interior of the tube for replacement; means for measuring the current flowing through the electrolyte as a function of time; An apparatus for determining the electrokinetic properties of an electrolyte in electrokinetic separation, comprising at least one means for determining an electroosmotic velocity, and means for measuring the velocity of a component inside a sample. 8. The apparatus of claim 7, wherein one of said plurality of electrolytes comprises a micelle. 9. The method of claim 1, wherein the first of the plurality of electrolytes comprises a first buffer comprising two compounds, and the second of the plurality of electrolytes is substituted for the first of the plurality of electrolytes. 8. The apparatus according to claim 7, wherein the second buffer comprises a second buffer containing the same two compounds, the ratio of the two compounds in the second buffer being different from that of the first buffer. .