JPS5952979B2 - How to measure the electrophoretic mobility of particles - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a method for measuring the electrophoretic mobility of particles.

A voltage is applied to the liquid sample contained within the electrophoresis cell and the time required for a selected particle to travel a preselected distance between two set marks is desirably determined. The video image of the sample is recorded in a vidicon system and reproduced on a monitor, which is then observed under a microscope. In previous methods and devices, particles or a large number of five After a cell is captured and deflected by a vertical electric field in a fixed number of containers, its mass is determined and subjected to further investigation.The electrophoretic mobility of a particle is determined when the observed particle determined by the deflection experienced in the laminar flow. This conventional method and theta device are disadvantageous for various reasons, for example, the buffer and sample flows require high stability, while the total applied volume of the sample is so low that the ion buffers used are low and repeated investigations are almost impossible.05 According to yet another known method, the electrophoretic mobility of particles is determined by laser light diffusing through the particles. It is determined by the Doppler effect of the frequency of .

The cost of methods for analyzing diffuse laser light (such as multichannel analyzers) is considerable. Furthermore, a method is also known in which the electrophoretic mobility of each fine particle is obtained by time measurement. A voltage is applied to a liquid sample in an electrophoresis chamber, and the transition time required for a selected particle to travel a predetermined distance during microscopic observation of this sample is measured manually, e.g. with a stop watch. Ru. Disadvantageously, the main error contained in this measurement result is due to the operator. The main consequences of this will depend on the operator's judgment on the one hand in visual observation and manual time measurement and on the other hand in the selection of particles for measuring time. Therefore, in order to eliminate errors in these series of measurements, it is necessary to obtain an average value, which has the disadvantage that it is not only time consuming but also expensive.

The present invention therefore seeks to provide a method which eliminates the above-mentioned drawbacks, increases the accuracy of electrophoretic measurements and reduces the technical expense and time involved in such methods. Furthermore, the present invention seeks to provide a method for accurately time-measuring the electrophoretic mobility of each selected particle in the simplest possible manner, which eliminates the various drawbacks.

The present invention relates to a method for measuring the electrophoretic mobility of particles, which involves applying a voltage to a sample placed in an electrophoresis cell, observing the sample through a microscope, recording it using video technology, It consists of an arrangement that measures the time required for selected particles of a sample to travel a predetermined distance as viewed on a monitor, and an image scanning signal is created from the recording of the sample and passed through a filter and subsequent digital conversion. The image scanning signal after passing is logically connected to a plurality of pulses arriving simultaneously at the monitor by comparison with an adjustable standard voltage, one of the simultaneously arriving pulses being the image scanning signal. one of the delay line synchronization pulses 4 and the delayed image synchronization pulse, characterized in that the other two pulses are produced from the line synchronization pulses of the image scanning signal, with a delay from the image synchronization pulse of the signal; and a first signal created by a logical connection that occurs when the digital image scanning signals are sent simultaneously causes a counting operation that is stopped by a further signal created by a logical combination. be done.

That is, the counting operation is started and stopped by the logical connection of the digital image scanning signal of the video recording of the sample and the delay line video periodic pulse. Advantageously, after the start of the counting operation has been triggered by a delay line synchronization pulse, each delay line synchronization pulse is rejected so that a further delay line synchronization pulse occurs after the counting operation has stopped. . Furthermore, it is advantageous to be able to set the two delay times of each delay line synchronization pulse individually or simultaneously. Additionally, it is advantageous to be able to adjust the pulse frequency and delay time of the delayed image synchronization pulse. It is further advantageous for the monitor to be brightened by means of a signal produced from the image and a line synchronization pulse, and for the signal to appear as vertical and horizontal marked lines on the screen of the monitor. It is desirable that the count panores are digitized and numerical values obtained. The time delay from the video sync pulse induces a periodic signal that is transformed to appear as a square of variable surface on the monitor screen, forming an image section that is Serves as a base for measurement. That is, whenever electrophoretic particles of the sample to be analyzed move within this image section, the time it takes for one particle to travel from one vertical limit of the image section to the other vertical limit is measured. This time measurement is performed by starting and stopping a counting operation, and the count panores are digitized and calculated. Starting and stopping of the counting operation is performed by logical combinations and delayed video synchronization pulses are generated by the digital image scanning signal of the sample recording. These delay line synchronization pulses that trigger the counting operation are used to trigger the counting operation to ensure that the start and stop of the counting operation occur at both vertical limits of the image section of the monitor. later rejected. Particles of the sample contributing to the electrophoretic numerical calculation are selected by balancing standard voltages with respect to the digital image scanning signal. Next, embodiments of the present invention shown in schematic diagrams will be explained.

The videocon 1 is connected via a separation amplifier 2 and a modulator 3 to a monitor 4, and via the separation amplifier 2 to a ladder filter 5, and furthermore a comparator 6 provided with a standard voltage Ur.
It is connected to a first power of an AND (logical product) element 7 and a second power, which is connected to the output of an OR (logical sum) element 8 .

One input of OR element 8 is connected to the output of AND element 9, and the other input of OR element 8 is connected to the output of AND element 10. The other output of the separation amplifier 2 is connected via an amplitude separator 11 to a pulse distributor 12, the first output of which is connected to a multivibrator 13, and the output is connected to two other multivibrators 14, 15. It is connected. The other outputs of the pulse distributor 12 are connected to respective inputs of AND elements 9 and 10 via two multivibrators 16 and 17 in series. The output of the multivibrator]4 is connected to the second input of the AND element 9, the input of the modulator 3, the first input of the OR element 18, and the first input of the AND element]9, respectively. Similarly, the output of the multivibrator 15 is connected to the second input of the AND element]0, another input of the modulator 3, the second input of the OR element 18, and the first input of the AND element 20. It is being done. The output of multivibrator 17 is of course connected to the modulator, and the output of AND element 7 is connected to each input of AND elements 19 and 20. AND element 19 has its output connected to the reset input of flip-flop 2, while AND element 20 is connected to the reset input of flip-flop 22. The output of flip-flop 21 is connected to the third input of AND element 9, and the output of flip-flop 22 is connected to the third input of AND element 10. The OR element 18 and the AND element 7 are connected to each input of the AND element 23 at its output example, and the output of the AND element 23 is connected to the reset input of the flip-flop 24.
The set inputs of flip-flops 21, 22 and 24 are applied to the clock pulse input. The output of AND element 7 is connected to each input of two AND elements 25 and 26,
A second input of AND element 25 is connected to a first output of flip-flop 24, and a second input of AND element 26 is connected to a second output of flip-flop 24, respectively. The outputs of the AND elements 25 and 26 are the flip
It is guided to two inputs of a flop 27, the output of which is connected to the input of an AND element 28. AND element 2
A clock pulse oscillator 29 applies a signal to the second input signal 8, and the output of the AND element 28 is connected to a counter 30. The videocon 1 produces a recording of the liquid sample to be analyzed in which the particles are contained.

This recording is displayed on the screen of monitor 4. The videocon 1 further produces an image evaluation scanning signal and its S
The N ratio is improved with a ladder filter 5. This image scanning signal is digitized by comparison with the standard voltage Ur in a comparator, so that the digital image scanning signal reaches the AND element 7. Periodic pulses are induced into the amplitude separator 11 from the video signal separated by the separation stage 2;
Next, the pulse distributor 12 separates the pulse into an image period pulse (ISP) and a line synchronization pulse (LSP). This line synchronization pulse (LSP) is delayed by a multipipelator 13. The two subsequent multivibrators 14 and 15 each have a different delay time j and are connected in series with the multivibrator 13, so that their respective outputs constantly supply two delay period line pulses, The pulses cause the monitor 4 to be brightly scanned by the modulator 3 and are visible as two vertical marks on the monitor screen. The positions of these marks are either changed in common by changing the delay time of the multivibrator 13, or individually changed by changing the delay time of the multivibrator 14, ]5. These marks 7 define the distance on which the time measurements of particles in the sample to be analyzed are based. The image period pulse (ISP) at the output of the pulse distributor 12 is delayed twice by the multivibrators ]6 and ]7, so that the delay period image period pulse (ISP) is
X is provided to the output of the multivibrator]7.

The lead edge of this pulse is determined in time by multivibrator 16 and the travel edge of the pulse is determined by multivibrator 17. The delay 1SP creates a horizontal mark on the screen of the monitor 4 via the modulator 3 and adjusts the brightness of the monitor 4.
The width of this mark depends on the set delay time of the multivibrator]7. By varying the delay time by the multivibrator 16, the space between the upper limit line of the horizontal mark and the upper image limit of the monitor 4 is adjusted. Thus,
Three marks on the screen of the monitor 4 represent freely adjustable rectangular image sections that are utilized.

According to the inventive method for measuring the time of sample particles, the travel time required for a selected particle to pass through an image section between two vertical marks is measured. However, time measurements can only be made when the particles move within a horizontally restricted image section. Selection based on brightness and or size, e.g. where particles have to be electrophoretically evaluated, can be achieved using image synchronization pulses (ISP).
This is done by balancing the standard voltage Ur when it is digitized by the comparator 6. The illustrated arrangement is set in a standby position for measurements by a pulse at control input 1C by setting flip-flops 21, 22, 24.

It is carried out after the preparation of the analysis target from a technical point of view of the operation. In the standby position of this arrangement, the pulses are detected by delay 1SP, one of the two delay line synchronization pulses and the AND elements 7, 9, 10 and OR element 8 when the pulses of the digital ISP occur simultaneously.
is set to the output signal of AND element 7 by the connection of Feed to count 30. The next pulse produced under the above conditions reaching the j output of AND element 7 passes through OR element 18 and AND element 23 and resets flip-flop 24, which in turn resets AND element 26. After Flip Flop 27
Reset. The reset of flip-flop 27 blocks AND element 28 and stops the counting operation of counter 30. The results of the count are digitized and evaluated by a computer in relation to any time measurement. A new pulse at the control input IC places the array in standby position for the next measurement. The counting operation in counter 30 is performed by analysis. It is started when a selected particle in the sample passes one of two vertical marks. To prevent a break in the counting operation of the counter 30, when another particle of the same type in the sample passes the vertical mark and causes the start of the counting operation of the counter 30, a further delayed LSP of the respective delay time is applied. is blocked after the count operation starts. This is accomplished by connecting the output of AND element 7 to the outputs of multivibrators 14, 15 in subsequent flip-flops 21, 22 which are coupled to AND elements 19, 20 and 9, 10. be exposed. A pulse sent to the output of AND element 7 and causing the start of a counting operation resets flip-flop 21 via AND element 19 or flip-flop 22 via AND element 20. Which of the two flip-flops 21 and 22 is set, that is, which vertical marker on the screen of the monitor 4 the sample particles pass through depends on which output of the multivibrator 14 or 15 the delay LSP is set to. Determined by whether the count operation is triggered. Depending on which of flip-flops 21 or 22 is set, either AND element 9 or ]0 is programmed. That is, one of the periodic delay LSPs of the multivibrator 14, which triggers the counting operation and sets the flip-flops 21 or 22 and 27, triggers the counting operation of the counter 30 at the output of the AND element 7. This is because it is not possible to create a stop pulse for this purpose.

The counting operation is then stopped at one input of the two AND elements 9, 10, where one of the two flip-flops is unblocked, by the simultaneous appearance of the delayed image period pulse and the pulse of the digital image scanning signal. This is done only by the delayed LSP. The multivibrators 14, 15 are followed by a pulse waveform unit (not shown) for the pulse waveform of the delay line synchronization pulse.

[Brief explanation of the drawing]

The figure is a schematic diagram showing an embodiment of the present invention. 1...Videcon, 2...Separation amplifier,
3...Modulator, 4...Monitor 5.
...Ladder filter 1, 6...Comparator, 7...AND element, 8...OR element, 9, 10...And Motoko, 11...
Amplitude separator, 12... Pulse distributor, 13, 14, 15, 16, 17... Multivibrator, 18... OR element, 19, 20
・・・・・・AND element, 21, 22, 24, 27...
...Flip Flop, 23, 25, 26, 28
......AND element, 29...Clock pulse oscillator, 30...Year counter, Ur...
・Standard voltage, ISP... Image synchronization pulse,
LSP・・・Line synchronization pulse.

Claims (1)

[Claims] 1. Applying a voltage to a sample placed in an electrophoresis cell, observing the sample with a microscope, recording with video technology, displaying on a monitor, and causing selected particles of the sample to move a predetermined distance. The image scanning signal is created from a sample recording, digitally converted by a filter line and subsequent comparison with an adjustable standard voltage, and the image scanning signal reaches the monitor simultaneously. one of the pulses is created by the delay from the image sync pulse of the image scan signal, and the other two pulses are created from the line sync pulse of the image scan signal, and the delay line sync A first signal produced by the logical combination that occurs when one of the pulses, the delayed image synchronization pulse and the digital image scan signal are sent simultaneously, triggers a counting operation that triggers a further signal produced by the logical combination. A method for measuring the electrophoretic mobility of particles, characterized in that two different time delays of each time delay line synchronization pulse are set not only individually but also simultaneously.