JPS6148865B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an apparatus for measuring the electrophoretic velocity of visible particles, such as biological cells, suspended in a solution.

Several methods have already been proposed for measuring the electrophoretic velocity of visible suspended particles, but each method has an appropriate suspended particle density, and the measurement results will become unstable if the density is higher or lighter than that. The reliability of the measurement decreases. For example, in the grid method, images of floating particles are formed on a grid, and the intensity of the light transmitted through the grid changes each time the particle image crosses the grid lines due to particle electrophoresis, so waves of changes in light intensity due to multiple particle images are superimposed. The electrophoretic velocity distribution of the particles is calculated by frequency analysis from the photometric waveform of the light transmitted through the grating. However, if the particle concentration is too high, the amount of light reaching the observation area will decrease, and the increase in the number of particles will instead cause scattering. If the light intensity is weakened, it becomes impossible to calculate the electrophoretic velocity distribution of particles, and if the concentration is too low, statistical fluctuations will increase, resulting in different velocity distribution profiles for each measurement even for the same sample. Become. Furthermore, in a method based on the basic principle of tracking individual particles and measuring their electrophoretic velocity, it is appropriate for the suspended particle concentration to be much lower than in the above-mentioned method; When the electrophoretic velocity distribution is determined by comparing the particle shape with the concentration distribution shape after electrophoresis for a certain period of time, it is appropriate that the particle concentration be quite high.

On the other hand, the sample has a high particle concentration when it is prepared, and when it is injected into the measurement area of an electrophoresis measurement device with something like a syringe, the particle concentration in the same area is extremely high at the time of injection, but the particle concentration is very high. The particle concentration gradually decreases due to diffusion and sedimentation, and it cannot be said that electrophoretic measurements can be performed with a solution that maintains the desired particle concentration.

In the past, the person performing the measurement injected the sample into the electrophoresis measuring device and then took the measurement at an appropriate time, so the measurement was carried out at a time when the particle concentration suitable for each measurement method was reached. However, it was difficult to make reliable measurements.

The present invention, after injecting a sample into an electrophoresis device, detects when the suspended particle concentration has reached a level suitable for the measurement device, that is, the measurement results are most stable.
The purpose of the present invention is to provide a device that automatically performs measurements at that time. The present invention will be explained below with reference to Examples.

The figure shows an embodiment of the invention. 1 is an electrophoresis tube filled with a solution that suspends sample particles;
Electrodes are inserted at both ends (not shown), and a concentrated suspension of sample particles is injected from a sample injection port (not shown). The figure shows only the measurement area of the electrophoresis tube 1, and the injected sample particles diffuse into the measurement area shown by diffusion, sedimentation, and electrophoresis, and the particle concentration increases suddenly at first, and then The concentration gradually decreases. Reference numeral 2 denotes a projection lens system, which forms an image of floating particles in the measurement area on the light receiving surface of the image pickup tube 3. An illumination light source is disposed outside the direction perpendicular to the optical axis of the projection lens system 2, and the floating particle image is formed as a bright dot on a dark background. A video signal output from the image pickup tube 3 is inputted to a signal processing circuit 4 via a gate G, and the electrophoretic velocity of sample particles is detected in this circuit. The video signal output from the image pickup tube 3 is also input to the integrating circuit 5. The integrating circuit 5 consists of an integrating capacitor and a leakage resistor in parallel with the integrating capacitor, and its output indicates the average level of the video signal. Since the particle image is a bright spot, this average level is high when the concentration of sample particles is high, and has a constant monovalent function relationship with the concentration of sample particles. Therefore, by applying the output of the integrating circuit 5 to the comparators 6, 6' and comparing it with an appropriately set reference level, it is possible to detect a specific sample particle concentration state. Therefore, the sample particle concentration at which the most stable measurement result can be obtained is determined in advance through experiments, and the reference level is set so that a signal is output from the comparator 6 at that time. Since an appropriate particle concentration value has a range, two comparators, 6 and 6', are prepared, and the reference levels given to each are provided with a difference corresponding to this range. Comparator 6 gives a signal when the concentration is higher than 6'. As described above, the sample particle concentration in the measurement region is 0 immediately before sample injection, suddenly increases to a high concentration, and then gradually decreases.
Therefore, when the sample is injected, the comparator 6' first outputs a signal, after a short time the comparator 6 outputs a signal, and then the signal of the comparator 6 disappears (becomes low level), and after a while the signal of the comparator 6' disappears. Therefore, if the gate G is opened by the set output of the flip-flop 7, which is set at the fall of the high level signal of the comparator 6 and reset at the fall of the high level signal of the comparator 6', the particle concentration gradually decreases. A video signal is input to the signal processing circuit 4 only during the optimum particle concentration period, and highly reliable electrophoresis measurement is performed.

An example of the signal processing circuit 4 will be explained. The image pickup tube 3 is digitally scanned and horizontal coordinates (x
Coordinate) When the video signal is at a high level within a certain range of designation information, 1 is added to the designated address of the first memory using the y-coordinate designation information at that time as address designation information. This addressing information combines several pieces of y-coordinate designation information to designate one address. With this configuration, an area of a certain width is horizontally divided into several sections on the image plane, and particles within each section are detected. Therefore, if the particle concentration is diluted appropriately, one particle will be in one section, and the next particle will be in that section until it enters the section from the left end and exits from the right end by electrophoresis. You can prevent it from coming inside. In such a density state, each address in the memory increases by 1 during one vertical scan as long as there is a particle image in the section corresponding to that address. (must be less than one line). Therefore, each address of the first memory is set to 1.
Scan once for each vertical scan, and for an address whose number of memories does not increase in the two scans, add 1 to that address of the second memory as address designation information of the second memory, and The memory at that address in the first memory is reset to zero. At this time, the number of memories stored at that address in the first memory is the time required for the particle image to pass through one section from the left end to the right end, measured in units of vertical scanning periods, so it is stored in the second memory. When the addressing information is set to , the storage in the second memory when the operation is continued while the gate G is open will indicate the distribution of electrophoretic velocity.

The above-mentioned embodiment belongs to a method of tracking the movement of individual particle images, and the particle concentration is suitably quite low, and the appropriate concentration range is wide in the low concentration region. However, the present invention is not directly related to the method of measuring the electrophoretic velocity of particles, but is applicable to any method of measurement and is highly necessary.

Furthermore, methods for detecting the appropriate concentration of particles are not limited to those that process video signals from a camera tube, but can also be used, for example, to form a particle image of a certain area (individual particles do not need to be separated) on the light-receiving surface of a photometric element. Any method may be used, such as comparing the output signal of the light receiving element with a reference level using a comparator or measuring absorbance.

The present invention has the above-mentioned configuration, detects an appropriate particle concentration, and automatically performs measurement only within that concentration range, thereby stabilizing measurement results and eliminating individual differences among measurement personnel, thereby providing highly reliable electrical power. Electrophoresis measurement becomes possible.

[Brief explanation of the drawing]

The drawing is a block diagram showing the configuration of an apparatus according to an embodiment of the present invention. 1... Electrophoresis tube, 2... Projection lens system, 3...
...Image tube, 4...Signal processing circuit, 5...Integrator circuit, 6, 6'...Comparator, 7...Flip-flop, G...Gate.

Claims (1)

[Claims] 1. A particle concentration measuring means in the electrophoretic measurement area of the sample particle suspension, a means for comparing the output of the same means with a preset level, and a means for operating within a predetermined particle concentration range based on the output of the comparing means. An electrophoretic measuring device comprising controlled electrophoretic measuring means for sample particles.