KR100991652B1 - Diagnosis method, a device for carrying out said method and a serum preparation method - Google Patents

Abstract

The present invention is industrially employed in the use of medicine, in particular molecular physiologic methods, and in the instruments used to investigate (screen) and monitor (monitor) the course of treatment and to select in vitro pharmaceutical preparations for treatment. Medicines for the diagnosis of oncological diseases and dosages thereof, which can be applied. The present invention is directed to multi-factor diagnostics. The method of the present invention records Rayleigh irradiated scattering emitted by a biological fluid solution and includes Rayleigh scattering intensity, depolarization coefficient, mass and shape of scattering particles, intermolecular interaction coefficient, diffusion coefficient of diffusion, characteristic viscosity of solution and fluid of particles Consisting of measuring the mechanical radius. Physiological fluids are embodied in the form of serum, plasma and lymph. The diagnostic equipment of the present invention comprises two optical receivers 2 which are firmly fixed to the laser module 1 and the optical unit 3. The signal from the photoreceiver is routed through a coordination board arranged on the computer main circuit board to an analog-to-digital converter 4 board operating simultaneously to the board connected to the computer. Equipment operation control and regulation functions are performed by the control board 5, where the equipment is powered by the merged power supply 6.

Diagnostic method, serum production method, oncological disease

Description

Diagnostic method, equipment and serum production method for the implementation of the method {DIAGNOSIS METHOD, A DEVICE FOR CARRYING OUT SAID METHOD AND A SERUM PREPARATION METHOD}

The present invention relates to the field of medicine, i.e., the diagnosis of oncological diseases using molecular physiologic methods in particular.

Diagnostic methods for measuring the physical counts of biologically active liquids that can be used to determine the condition of patients by preparing serum are known (the Copyright certificate of SSSR No. 1319705, MPK A 61 V 10/00). This physical coefficient is measured by electronic box resonance (EPR).

The drawback of this method is that it requires expensive equipment to implement it and the implementation is complex.

Another well-known diagnostic method is the physical count of Rayleigh scattering of laser irradiation by a solution of serum to prepare serum and its solution, to affect the serum solution by irradiating the laser and to determine the condition of the patient. And measuring them (Patent RF No. 2000027, MPK G 01 N 33/483). In this method the serum is pre-dissolved with distilled water. The resulting solution refers to a biologically active solution. Rayleigh scattering rate, effective weight and intermolecular interaction rate of scattering particles are measured as physical coefficients.

The drawback of this method is that it only measures the average weight of the particles scattered upon implementation of the method.

Most known diagnostic methods include preparing a biological liquid solution, irradiating a laser to affect the solution, and measuring the physical coefficient of irradiation emitted by the solution of the biological liquid to determine the patient's condition, Also in the measurement of the physical coefficient of radiation emitted by a solution of biological liquid, the signal is analog-to-digital converted and computerized (the Patent of the Russian Federation No.2219549, MPK G 01 N 33/52). In this method, the physical coefficient of weak water solution of the patient's natural serum is measured by laser correlation spectroscopy. Two solutions are prepared for this purpose, in which one is added an alkali and the other is added an acid. The possible density of the variable amplification distribution of the light dispersion intensity in each of these solutions is estimated.

The drawback of the diagnostic method most similar to the technique of the present invention is that the number of adjustable coefficients is not so large when performing the diagnosis.

The device most similar to the device of the invention comprises a laser module with a laser radiation source, an optical unit with a cuvette unit, a light receiving module comprising at least one analog-to-digital converter, the cuvette unit Optically connected to the optical receiver block and the analog-to-digital electrical converter is electrically connected to the optical receiver and the computer (the Patent of the Russian Federation No. 2219549, MPK G 01 N 33/52).

This instrument is used to measure the physical coefficients of weak aqueous solutions of the patient's natural serum by laser correlation spectroscopy. Two solutions are prepared for this purpose, one of which is added an alkali and the other an acid. The possible density of the variable amplification distribution of the light dispersion intensity in each of these solutions is estimated.

The drawback of the most similar equipment is that the number of adjustable coefficients is not very high when performing a diagnosis.

Known methods for preparing serum include taking natural blood from a fasting patient and placing it in a sealable test tube (the Patent of the Russian Federation No. 221949, MPK G 01 N 33/52).

The drawback of this similar method is that the conditions for preparing the serum are not optimal, which reduces the reliability of the diagnosis.

The present invention solves the technical problem of realizing multi-factor diagnostics.

An object of the present invention is obtained by a diagnostic method comprising preparing a biological liquid solution, irradiating a laser to the solution and measuring the physical coefficient of radiation emitted by the biological liquid solution to determine the condition of the patient, In the measurement of the physical coefficient of irradiation emitted by the biological liquid solution, the signal is converted into an analog-to-digital converter and processed by a computer. As the biological liquid, the present invention uses at least one of the following liquids: serum, plasma and lymph.

It also measures one or more of the molecular coefficients including Rayleigh scattering intensity, depolarization coefficient, weight and shape of the scattering particles, intermolecular interaction factors, transfer diffusion coefficient, characteristic viscosity of the solution, and hydrodynamic radius of the particles.

In particular, it is possible to measure the molecular count of at least one protein. Therefore, it is possible to determine the molecular count of at least one protein from the group consisting of: albumin, α'-globulin, α'1-globulin, β-globulin and γ-globulin.

In particular, it is possible to measure the physical coefficient of fluorescence irradiation emitted by a biological liquid solution.

In particular, it is possible to adsorb a compound which fluoresces in the visible region of the spectrum on the surface of the molecule of the protein.

In particular, it is possible to measure the beating spectrum between Rayleigh dispersion irradiation of a biological liquid solution and basic determined laser irradiation. At this time, the laser irradiation and the basic measured laser irradiation can be obtained from the same source. In addition, the laser irradiation affecting the biological liquid solution and the basic measured laser irradiation can be aligned with each other in the 90 ° direction. In addition, Rayleigh dispersion irradiation can be measured in the dispersion direction of the basic measured laser irradiation. Therefore, Rayleigh dispersion irradiation can be measured by being aligned with each other by 180 ° in two directions.

In particular, in the measurement of the molecular count of the radiation released by a biological liquid solution, it is possible to change the pH of the solution. In particular in the measurement of the molecular count of the radiation released by the biological liquid solution, it is possible to change the concentration of the solution.

In particular, patients with negative intermolecular interaction coefficients between serum and lymph may be classified as tumor patients.

In particular, the correlation function of the irradiation scattered on the particles of the solution can be estimated.

In particular, it is possible to estimate a function of the spectral density of the radiation scattered on the particles of the solution. At this time, it is possible to measure the relative magnitude of the two consecutive spectral maximums. In this case, the relative magnitude of the half-width of the two consecutive spectral maximums can be determined.

In particular, the volume of the biological solution may not exceed 1 ml of liquid.

In particular, it is possible to filter the signal before the analog-to-digital conversion of the signal.

An object of the present invention is a diagnostic means comprising a laser module comprising a source of laser irradiation, an optical unit comprising a cuvette unit, at least one optical receiver and an analog-to-digital converter, the cuvette unit being optically connected with a module of the optical receiver. The analog-to-digital electric converter is electrically connected to the optical receiver and the computer, the optical block additionally includes a light splitting chevron connected to the laser irradiation source and the cuvette unit, and the laser module and the optical receiver module are connected to the optical unit. It is firmly fixed.

In particular, the light splitting chevron is firmly fixed to the optical unit.

In particular, the equipment may further comprise a control board electrically connected with the laser module and programmed to perform the commissioning and deactivation tasks of the laser module.

In particular, the optical unit may further comprise a unit of optical circuit formation optically connected to the cuvette unit and securely fixed to the optical unit.

In particular, the photoreceiver module may further comprise an optical inlet forming block, which is optically connected with the photodetection equipment and securely fixed to the module.

In particular, the equipment may further comprise a low-noise amplifier electrically connected with the module of the optical receiver.

In particular, the equipment may further comprise a standardized amplifier electrically connected to the low-noise amplifier.

In particular, the analog-to-digital converter may further include a timer-set filter and buffer memory installed on one board.

In addition, an object of the present invention is a method for preparing plasma, which method takes blood from a patient on an empty stomach, puts it in a sealed test tube within 60 minutes, and puts it in a thermostat at 35-40 ° C. for 20 to 120 minutes, The test tube is then cooled to a temperature of 1 to 10 ° C., maintained at this temperature for 10 to 90 minutes, and further centrifuge the test tube with blood at 1000 to 4000 g for 10 to 60 minutes (where g is gravity Acceleration), taking one to three ml of blood from a sealed test tube after centrifugation.

In particular, it is possible to take 0.1-100 ml of blood from the patient.

In particular, the blood can be taken from the patient without a special device.

In particular, blood may be filled with at least 60% of the sealed test tube volume.

In particular, the blood may be placed in one of the sealable test tubes during the loading of the blood.

In particular, 5 to 20 test tubes can be placed on the thermostat at the same time.

In particular, the test tube can be cooled in a water bath.

In particular, the cooled test tube can maintain 1 ~ 10 ℃ by putting the test tube in the refrigerator.

In particular, the test tube may be sealed around the rim after the blood.

In particular, 4 to 20 test tubes containing blood can be centrifuged at the same time.

In particular, clear serum can be taken from above the coagulum formed by centrifugation in the center of the test tube.

In particular, the serum may comprise at least 90% of the cuvette dose.

The present invention provides a diagnostic method, wherein the equipment for realizing the diagnosis and the method for producing the substance to be measured have a certain correlation with the present invention.

In the use of the present invention, the volume of serum or plasma for analysis is sufficient to 1 ml, and can be made sufficiently from the blood remaining for use for standard biochemical comprehensive medical examination. Average analysis time takes 3-15 minutes depending on the speed of your computer. The present invention enables large-scale diagnosis at high speed, and thus can diagnose early stage tumor patients which are hardly found by conventional methods of diagnosis. Thus, it is advantageous to treat patients in risk groups with a tendency to develop new tumor growth. The present invention can be used to adjust the treatment efficiency for the application of all modern tumor treatment methods and to select in vitro an effective medicament in a model of a patient's serum or blood plasma. The present invention makes it easy to individually control the treatment of a patient.

The development of pathological phenomena in body tissues is accompanied by changes in some molecular counts in cells, proteins, and plasma. Serum contains proteins of varying weight and proportion.

Serum base protein is albumin weight 70000 (50%), α'-globulin, average weight 40000 (6%), α'1 globulin, average weight 50000 (11%), β-globulin, average weight 70000 (7%) γ-globulin is considered to be 150000 (16%). The average molecular weight of the serum particles is estimated at 100000.

Protein molecules have a charge on their surface, which results in large dipole protein momentum of several hundred D (debye). The surface charge of protein molecules in solution can vary over a wide range depending on the concentration change ��� of the free protons (ie pH). Because of this property, there is a strong electrostatic interaction between the charges of the protein molecules and between the polar solvent and the charged surface groups of the protein, which affect the properties of the Brownian motion of the molecule.

The dispersion of light in serum and lymph is determined by the physical count of dissolved molecules. Aqueous protein solutions can be used as a model for biological liquids such as blood or lymph.

Rayleigh dispersion of light is the most direct and effective way to study the intermolecular interaction, mobility and polarity of macromolecular solutions, including aqueous protein solutions. This method can be used to directly determine the molecular weight M, for which the Rayleigh scattering factor or turbidity R90 is measured at various concentrations (C) of the solution and the dependence measured by extending the size CH / R90 as a function of concentration Extrapolation to C = 0 is necessary. The slope of this straight line, 2B, allows the calculation of the second Viral coefficient, which serves to characterize the degree of deviation of the behavioral properties of the solution from the ideal solution and to measure the intermolecular interactions in the solution.

In the case of solution of protein charged molecules, the effect of surface charge on the surface of molecules and the behavior of molecules in solution, in particular on the coefficient of interaction between molecules, is very large. If the surface charge on the molecule of a protein changes, there is a possibility of the development or growth of a tumor disease as a result. As the charge decreases, the Coulomb force that pushes the intermolecular proteins weakens and the attraction between them becomes stronger. This changes the magnitude and sign of the coefficient B and forms a complex of molecules with a higher molecular weight than each protein.

The dynamic coefficients of Rayleigh dispersion of light are estimated using Fourier photon analysis of the spectral components of diffuse light. In this method, the correlation function of the variation of the diffused light intensity due to the Brownian motion of the solution particles is studied. At this time, the particle diffusion coefficient and their hydrodynamic radius can be measured. The concentration dependence of the diffusion and dispersion coefficients is defined by the same Viral coefficients in the factor. The kinetic coefficients of charged molecules in solution depend essentially on the surface charge of the protein molecules. When a tumor-related disease is present, a change in the surface charge of the protein molecules in the blood occurs, which results in a change in the density of the spectral components of the diffuse light and the half-width of the spectral components.

As tumor-related diseases progress, early stages of autocatalytic reactions involve the formation of free radical states in cells and protein tissues. This process can be easily found by fluorescence analysis using the corresponding probe. An early stage of aggregation of serum proteins can be found by examining the polarized fluorescence of various probes associated with proteins in the serum, including chelates.

When a fluorescent substance containing a solution of protein molecules is irradiated with monochromatic linearly polarized light, the anisotropy of fluorescence of solution particles occurs. Rotational fusion, which occurs during the excited state and changes the emission direction of the dipole phosphor, can reduce the measurable anisotropy magnitude below the maximum value.

The fluorescence of a protein is defined by its amino acid structure and observed in the ultraviolet region. Therefore, when studying proteins in aqueous solution, compounds are specifically selected that adsorb on the surface of protein molecules and fluoresce in the visible part of the spectrum. As such compounds, fluorescent probes or dyes are used. Probes with large quantum quantities fluoresce strongly in the visible region and obtain information about the molecular mobility of proteins from these fluorescence coefficients. By means of probes it is possible to study the surface charge of molecules of proteins.

The invention is further illustrated by the detailed description of the accompanying drawings and the experiments to be performed on the drawings.

1 is a schematic diagram of the equipment of the present invention.

2 is a schematic diagram of a laser module.

3 is a schematic view of an optical unit.

4 is a schematic diagram of an optical receiver module.

5 shows a single-parameter histogram of factors of intermolecular interaction in group 3 patients (health group, risk group and tumor patient group).

Figure 6 shows the dependence on the pH size of the aqueous solution BSA and γ-globulin of the transfer diffusion coefficient.

7 shows the results of a clinical investigation of two coefficients of health and patient groups.

The diagnostic equipment (FIG. 1) comprises a laser module 1 and two optical receivers 2 modules firmly fixed to the optical unit 3. The signal from the optical receiver 2 enters the analog-to-digital converter 4 board, which is simultaneously connected to the computer via a coordination board mounted on the main circuit board of the computer. Function control and function maintenance of the equipment is made by the control board 5, and the electric supply thereof is performed by the power supply 6 installed therein.

The laser module 1 (FIG. 2) consists of a laser diode 7 and its power supply and controller 8, which keeps the light emission of the laser diode 7 stable and from the control board 5. Adjust the operation according to the signal.

The module 2 (FIG. 3) of the optical receiver serves to detect the Rayleigh distributed signal and to elevate the signal to the level necessary to supply the signal to the analog-to-digital converter 4. It consists of an optical inlet forming block 9, an optical receiver 10, a low-noise amplifier 11 and a standardized amplifier 12. The optical inlet formation block 9 stably forms the wavelength of the front portion of the scattered irradiation, which provides the optical receiver 10 with the maximum useful signal. The low-noise amplifier 11 maximizes the required signal by minimizing as many as possible noise characteristics. The normalization amplifier 12 is a means for providing a signal at a value that provides an optimal use of the dynamic range of the analog-to-digital converter 4.

The optical unit 1 (FIG. 4) consists of a light splitting chevron 13, a cuvette unit 14 and an optical path forming unit 15. As shown in FIG. The cuvette unit 14 fixes the cuvette containing the probe fluid having the accuracy necessary for the beam of the laser diode and the entrance aperture of the optical receiver. The optical path forming unit 15 provides the adjustment and fixing of the position of the laser diode 7, the light receiver 10 and the optical forming elements during the measurement. The optical path forming unit 15 has a strictly adjusted structure in which the diaphragm, the adjusting unit and the immobilizing elements are installed.

The analog-to-digital converter 4 limits (filters) fragments of the frequency of the analyzed signal, converts the filtered analyzed signal from analog to digital, preserves the buffer memory before writing the result to working memory, and analyzes it. It is designed to control the digital accumulation of the signal and to record the transmission of computer signals through the coordination board installed in the ISA slot of the main circuit board. The analog-to-digital converter 4 board is made in the form of a printed circuit board and is separated for connection with a computer, and has two inlets that accept analog signals and can store 1048 samples at a time. The number of analog-to-digital converters is twelve. The analog-to-digital converter (4) board can be operated in data accumulation mode and data transfer mode. The converter board consists of an adjustable timer, buffer memory, a device for converting the signal to digital, and a buffer register that stores the upper or lower byte counts for management to send its report to the computer.

The control board 5 allows the equipment to operate properly in normal and emergency modes. This control board allows the equipment to be turned on and off in a certain pattern, to determine whether the equipment can work normally, to control the efficiency of the functional blocks of the equipment, to control the access and acquisition of information about the status of the equipment, For those things, it tells them to do things like connect with an external computer running through a series of ports.

The power unit 6 is for supplying a constant coefficient of voltage which provides optimum performance of the equipment for a frequency of 50 Hz and an inlet power of 220V.

The principle of operation of the equipment (FIG. 1) is based on laser correlation spectroscopy, which consists in measuring the spectra of the quasi-elastic capacity of diffuse light. It is known that the dispersion of light on the particles in the Brownian particles expands the width of the initial emission spectrum, i.e., leads to diffusion. Since the width of the laser irradiation spectrum is very narrow, the width of the spectrum of diffused light is proportional to the diffusion coefficient of analysis associated with the size of the scattering particles analytically according to the known Stokes-Einstein equation.

In the case of a polydispersion system, if diffusion is caused by dispersion of particles having various diffusion coefficients, the problem of defining the scattering particle distribution function according to the size leads to complex mathematical processing of the experimental spectrum. Equipment to solve this problem is achieved by mathematical standardization. After standardization of the function of determining the scattering particles, they are input into a computer database for statistical analysis of various coefficients.

After taking 5 ml of blood from a fasting patient (preferably by drift), the taken blood was placed in a sealed test tube, and the volume between the surface of the blood and the stopper was 1 ml. A set of 10 test tubes was placed in a thermostat at 37 ° C. for 60 minutes.

After removing the test tube from the thermostat, the test tube was rounded, rounded, and each test tube was again sealed.

Then, the test tube containing the blood was placed in water of 5 ℃ and transferred to the refrigerator of the same temperature and stored for 60 minutes. Then, a set of test tubes were taken out of the refrigerator and everything was rounded. A set of test tubes was placed in a centrifuge. After centrifugation at 15000 g for 30 minutes, lumps formed in each test tube. After centrifugation, serum was taken from the center of the test tube (on lumps) using an eyedropper and placed in a sealable 1.5 ml cuvette to minimize the volume of air between the surface of the serum and the stopper. When taking serum, it is important not to touch the mass. The selected serum should be visually examined, which should not contain suspended solids, jellyfish-like object formation and surface membranes. If the serum does not meet this requirement, it is not appropriate for the test. A sealed set of cuvettes containing serum was stored in a 5 ° C. refrigerator until tested. Four blood tests were performed at the same time. The time from taking the blood to testing the blood should not exceed eight hours.

Equipment for analyzing scattering in serum in aqueous solutions using correlation spectroscopy, including the analysis of the integration and dynamic dispersion of diffused light, using the investigation of coherent excited states, can be used for static and dynamic molecular counts (weight, morphology, intermolecular interactions). Coefficients, depolarization coefficients, and diffusion coefficients, characteristic viscosities, and hydrodynamic radii) provide multi-coefficient diagnosis for a wide range of diseases, particularly tumors.

Unlike similar prior equipment, the present invention allows simultaneous measurement of the static and dynamic coefficients of particles (proteins or their aggregates) in aqueous solutions of serum. Therefore, the course of treatment can be examined (screened), monitored (monitored), and screened for in-vitro drugs and the suitability of the doses for their administration.

The Rayleigh light scattering method of the present invention is used for the diagnosis of cancer. The relative intensity of Rayleigh dispersion R (90 ° to the falling beam), the weight M of scattering particles and the intermolecular interaction coefficient B were measured. All three coefficients for serum solutions in healthy and tumor patients show a marked difference (see Table 1).

Table 1 shows the B values for healthy people and tumor patients. In the case of tumor patients, the B value changed significantly and became negative, and the weight of the scattering molecules increased. For patients without tumors, B and M values were little different from those of healthy patients. In FIG. 5, histograms of intermolecular interaction B-value factors for health group 16, risk group 17, and tumor patient group 18 are shown. The histograms for the healthy group 16 and the tumor patient group 18 did not substantially overlap (FIG. 5).

Using the Malvern correlator for protein (19) and γ-globulin (20), the dynamic diffusion coefficient Dt, detected by dynamic light scattering, changes with the surface charge in the nonlinear phase molecules with the lowest isoelectric point ( 6).

The clinical findings of the health group 21 differed in two coefficients representing the magnitude P of the concentration of albumin and globulin and the width W of the spectral alumina in the risk group 22 and the patient group 23 (FIG. 7).

Several naphthylaminesulfonic acid fluorophores were used as probes in the use of fluorometry. The probe is not covalently bound to the protein and hardly fluoresces in water, but strongly fluoresces when bound to the protein molecule. Weak quantum emission in the water phase means that the fluorescence irradiation portion does not have to be taken into account. The degree of anisotropy or polarization of fluorescence is also a diagnostic factor and increases the ease of diagnosis.

The present invention enables industrial application of equipment used to examine (screen) the course of treatment and monitor (monitor) the course of treatment, and to select agents and their dosages in vitro for treatment.

Claims (39)

delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete A laser module 1 with a laser irradiation source 7, an optical module 3 with a cuvette unit 14, a module of at least one optical receiver 2 and an analog-to-digital converter 4, the cuvette unit 14 is optically connected with the optical receiver 2, and the analog-to-digital electric converter is a diagnostic equipment electrically connected with the optical receiver 10 and the computer, and the optical unit 3 is further provided with laser irradiation (7). A light splitting chevron 13 optically connected to the source and cuvette unit 14, wherein the laser module 1 and the optical receiver module 2 of the optical receiver 2 are securely fixed to the optical unit. equipment. The diagnostic device according to claim 20, wherein the light splitting chevron (13) is firmly fixed to the optical unit (3). 21. The diagnostic device according to claim 20, wherein the equipment further comprises a control board electrically connected with the laser module (1) and performing a task to cause the laser diode (7) to be turned on and off in a predetermined pattern. 21. Diagnostic equipment according to claim 20, wherein the optical unit (3) further comprises an optical path forming unit (15) optically connected to the cuvette unit (14) and rigidly fixed to the optical unit (3). 21. Diagnostic equipment according to claim 20, wherein the module (2) of the optical receiver further comprises an optical inlet forming block (9) optically connected to the optical receiver (10) and rigidly fixed to the module. The diagnostic device according to claim 20, wherein the device further comprises a low noise amplifier (11) electrically connected to a module of the optical receiver (2). 21. The diagnostic device of claim 20, wherein the device further comprises a standardized amplifier (12) electrically connected with the low-noise amplifier (11). The diagnostic device according to claim 20, wherein the analog-to-digital converter (4) comprises a filter with a timer set and a buffer memory mounted on one board. Place the sealable test tubes in a thermostat at 35-40 ° C. for 20-120 minutes, before placing the blood in one or more sealable test tubes and after 60 minutes after placing the blood in the sealable test tubes, and then place the test tubes in 1 Cool to a temperature of ˜10 ° C., maintain this temperature for 10-90 minutes, further centrifuge the test tube with blood at 1000-4000 g for 10-60 minutes, and 0.1-3 ml from the sealed test tube after centrifugation Method for producing a serum comprising extracting the serum of. 29. The method of claim 28, wherein the volume of blood is 0.1-100 ml. delete The method of claim 28, wherein the blood comprises at least 60% of the volume of the sealable test tube. delete 29. The method of claim 28, comprising simultaneously placing 5 to 20 test tubes in a thermostat. 29. The method of claim 28, comprising placing the test tubes in a container containing water to cool the test tubes. The method according to claim 28, wherein the test tube is kept in a refrigerator to maintain 1 to 10 ° C. The method of claim 28, wherein the test tube is rounded and then sealed. 29. The method according to claim 28, wherein 4 to 20 test tubes containing blood are centrifuged at the same time. 29. The method of claim 28, comprising taking transparent serum over the coagulum formed by centrifugation from the center of the test tube. The method of claim 28, wherein the blood comprises at least 90% of the cuvette volume.

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