Average Volume Digital Computer And Digital Volume Totalizer For Cells And Particles

Abstract

An apparatus for deriving average volume and total volume data from particles in a flow of liquid metered through an orifice includes a display of mean particle volume and total particle volume and count. The mean particle volume is the quotient of the totalized particle volume of a specific number of particles divided by that same number. The total particle volume and count provided by the same apparatus can be used to confirm the mean particle volume measurement.

Description

BACKGROUND OF THE INVENTION

It has long been the practice in clinical laboratories to determine the fraction of cell volume in whole blood by centrifuging blood in a thin, constant diameter tube, and then manually positioning the tube adjacent to suitable scale to read the "hematocrit" percentage at the interface between the dark red packed cells and the comparatively clear plasma. This procedure is primarily manually performed and is, therefore, a time consuming operation.

Another method for determining the percent of cell volume involves the use of an orifice tube and orifice current supply in combination with vacuum apparatus well known in the art to generate electrical pulses which are proportional to particle volume. The pulses are operated upon, for example, by shaping to provide an integrated charge on a capacitor which is utilized to operate a motor mechanism for a scale pointer, pen recorder or the like. Such procedure, however, suffers from changes in ambient conditions and analog component stability and requires apparatus for shaping the received pulses.

SUMMARY OF THE INVENTION

The primary object of the present invention is to overcome the drawbacks of the aforementioned types of particle volume measuring procedures.

Another object of this invention is to provide directly for the case of blood analysis an improved method for measuring not only the percentage of cell volume in whole blood (HCT) but the average cell volume (MCV), which may also be obtained by dividing the hematocrit percentage (HCT) by the red blood cell (RBC) count in a unit volume of blood and multiplying by a power of 10.

Another object of this invention is to provide the precision and stability of digitally computed parameters in a manner providing double checking, for example, by confirming the MCV by dividing the HCT by the RBC count and multiplying by a power of 10, and accomplishing the foregoing without resort to a programmed digital computer with the attendant cost of complex, sophisticated circuitry and software.

A further object of this invention is to provide relatively simple, accurate instrumentation for widely used applications which require this type of particle volume analysis for large numbers of samples, such as clinical laboratories for red blood cells.

An instrument apparatus, for example, scans a flow of 2,560,000 cubic microns of whole blood in a diluted state to generate a pulse for each cell traversing an orifice such that the pulse amplitude is directly proportional to the volume of the cell. These pulses are suitably factored to provide the direct RBC count.

For determining the HCT value, these pulses are also presented to an analog to digital converted (ADC) which provides a digital pulse train output for each cell such that each pulse of said train represents two cubic microns of cell volume. This pulse train from the ADC is then divided by 12,800 for direct display on a counter as the HCT percentage when the RBC count is completed. The apparatus includes means for producing start and stop signals for these displays which correspond to the initiation and termination of the scanned flow through the orifice, in addition to means for producing the individual particle pulses, for converting the amplitudes of said pulses to a digital pulse train, and for properly dividing the pulse train.

The apparatus, according to the present invention, is also provided with independent means for measuring and displaying the MCV directly in cubic microns, by which the digital pulse train output of the ADC is divided by a suitable factor 2,048 being an exemplary illustration, and this divided output is counted until stopped by a signal provided at a count of twice that number of cells, i.e., 4,096 cells. Again, each pulse of said train means two cubic microns of cell volume; the total volume of the 4,096 cells thus being divided by 4,096 in an "on-line" manner.

BRIEF DESCRIPTION OF THE DRAWINGS

Other objects, features and advantages of the invention, its organization, construction and operation will be best understood from the following detailed description of an exemplary embodiment thereof, taken in conjunction with the accompanying drawings, in which:

FIG. 1 is a circuit block diagram of particle volume analyzing apparatus according to the invention;

FIG. 2 is a circuit diagram of a pulse analog to digital converter which may be employed in the circuit illustrated in FIG. 1;

FIG. 3 is a graphical illustration of wave forms and event sequences at selected points in the circuit illustrated in FIG. 2;

FIG. 4 is a circuit representation of an accumulator which may be employed in the circuit of FIG. 1; and

FIG. 5 is a graphical illustration of wave forms and event sequences at selected points in the circuit illustrated in FIG. 4.

DESCRIPTION OF THE PREFERRED EMBODIMENT

An electric sensing zone 10 comprises a container 11 holding therein conductive liquid 12 containing particles to be analyzed and an agitator 15 operable to maintain the particles continuously suspended in the liquid 12. An orifice tube 13 which is also filled with the liquid 12 is disposed in the container 11 and includes an orifice 14 to pass the liquid into the tube 13 in order to establish an electrical circuit between an electrode 22 within the tube 13 and an electrode 23 disposed below the surface of the liquid 12 in the container 11. The tube 13 includes an end for connection to a source of negative pressure, a vaccuum supply, the application of such negative pressure being controlled by an interposed valve 19.

Also connectable to the source of vacuum by way of the valve 19 is a mercury siphon tube 16 having an open end 18 and containing a supply of mercury. Located within said siphon tube 16 is a pair of electrodes 20, 21 for controlling the start and stop operations of particle and total volume count, as will be explained in greater detail below.

The electrodes 22, 23 are connected by way of a pair of conductors to a current supply 24 to establish a current flow in the circuit including the electrodes 22, 23 as is well known in the art. Further, and as also known in the prior art, the passage of particles through the orifice 15 modulates the current flow during their passage to produce a sequence of voltage pulses. These pulses and their amplitudes represent the particle population and size of particles in the liquid 12.

The particle pulses are fed to a linear pulse amplifier 25 which has an output connected to an adjustable trigger circuit 26 for triggering a count register 27 during the interval of engagement of the mercury 17 with the start and stop electrodes 20, 21, respectively. Reference may be had to U.S. Pat. No. 3,345,502 for a more detailed discussion of this type of apparatus and its operation which will not be treated in further detail herein. It suffices here to state only that the amplifier 25 is provided with particle pulses whose amplitude is directly proportional to particle volume and that there is an interval of pulse count corresponding to the contacting of the mercury with the electrodes 20, 21 after the valve 19 to the regulated vacuum has been closed.

The linear particle pulse amplifier 25 provides the particle pulses to a pulse amplitude digitizer 29 (ADC) which converts the amplitude of the particle pulses into digital information signals. This digital information is made available to an accumulator which operates as a particle pulse summator in an overflow mode.

The accumulator 30 has a first output tapped off at an intermediate count which represents in digital form the total volume of the particles which have traversed the orifice 14. This information is provided to a total count volume register 31 which includes apparatus for displaying the total volume. A second output of the accumulator is connected to a mean volume register 32 which is effective to register the mean particle volume by summing overflows of the accumulator. As can be seen from the drawing, the total count volume register is operable for a period of time governed by the start and stop electrodes 20, 21; whereas the mean volume register has its start control governed by the start electrode 20 but its stop control provided by way of a predetermined count stop apparatus 33 which is driven by the trigger circuit 26. The predetermined count apparatus 33 is provided as a factor arrangement whereby the predetermined count may be evenly related to the volume of particles on a simple decimal multiple basis of the average particle volume.

In a particular system constructed which is capable of running a typical RBC operation, the predetermined count from the apparatus 30 was selected to be 4,096 and the overflow count was selected to occur at 2,048 in that the apparatus provides two counts per cubic micron. The output to the total count volume register 31 was selected to occur at each count of 128, this count being divided by 100 in the total count volume register. Therefore, 128 pulses equals 256 cubic microns and their summation divided by 100 yields pulses meaning 25,600 microns. With blood diluted at 62,500 .times. 160 microliters of such a dilution equals the aforementioned 2,560,000 cubic microns as a sample of whole blood to be scanned. While reference is made herein to conducting an RBC operation, it is clearly evident that the instant invention is applicable for analyzing many other types of particle populations.

Referring to FIGS. 2 and 3, the pulse amplitude digitizer 29 and its operation will be set forth. It should be noted that the lower case reference characters a- n adjacent wave shapes in FIGS. 2 and 4 correspond to the trace references in FIGS. 3 and 5 and arrows of the reference characters 0- t are influences or cause and effect references. The particle pulses a are received by the digitizer 29 on an input conductor 34 and fed to an operational amplifier stage which is constructed for unity gain. The operational amplifier stage includes an amplifier 35 having the conductor 34 connected to its positive input, a diode 36 serially connected to the output of the amplifier 35 and a feed-back conductor 37 connecting the diode 36 to the negative input of the amplifier 35. A capacitor 38 is connected to the diode 36 and is effective to store energy in response to a particle pulse, the voltage across the capacitor representing the amplitude of the particle pulse, and is effective to remember the pulse amplitude. The pulses received on the conductor 44 are also connected to the positive input of an amplifier 40 which has its negative input connected to a variable resistor 41 which is adjustable to set a threshold level or noise discrimination level a' so that the apparatus is rendered insensitive to low level signals. The amplifier 40 operates to provide an output pulse having a width determined by the crossings of the level a' by a particle pulse. This output signal c is connected to the set input of a flip-flop 42 which is responsive to initiate an output pulse d for turning on a constant current generator device 43 which includes a switch 44 and a constant current generator 45. The activation of the constant current apparatus 43 is effective to discharge the capacitor 38 in accordance with the expression T = C V/I, where I is the constant current, C is the capacitance of the capacitor 38, V is the initial voltage across a capacitor 38 and T is the discharge time of the capacitor 38. With the capacitance and the current being constant, the discharge time is shown to be linear. The resulting wave form b is applied to the positive input of an amplifier 39 to terminate a pulse e which was initiated in response to the initial start of charging of the capacitor 38 as the voltage thereacross went above zero.

The pulse e is fed to a differentiator circuit 46 which responds to the trailing edge of the pulse e to provide a spike f for application to the reset input of the flip-flop 42 which causes the flip-flop to reset and termination of the pulse d.

The output of the flip-flop 42 is also applied to one input of a AND gate 47 which has another input connected to a clock (pulse generator) 48. The pulse d is effective to gate the output of the clock through the gate 47 to a gate 49 which passes the digitized output signal h. The pulse d itself is likewise passed through a gate 53 as an available invented output signal d.

In order to reject high amplitude pulses which are not within the range of sizes of particles being evaluated, for example a fiber in a blood sample, an upper limit threshold level a is established by means of a variable resistance 51 connected to the negative input of an amplifier 50. The positive input of the amplifier 50 is connected in common with the positive input of the amplifier 40 to the input conductor 34. The amplifier 50 has its output connected to a flip-flop 52. The amplifier 50 produces a pulse j having a width determined by the crossing of the upper threshold a by a particle pulse. The pulse j is effective to set the flip-flop 52 and initiate a pulse k' which is terminated upon reset of the flip-flop 52 by way of a pulse f which was initiated in response to the discharge of the capacitor 38 as described above. The pulse k' is effective to block the gates 53 and 49 and prevent outputting of the respective signals d', h therefrom.

Referring now to FIGS. 4 and 5, the accumulator 30 and its relation to the digitizer 29 and to the volume display register 31 and 32 is illustrated. The output h is supplied by way of a conductor 54 to a counter 55 which divides the output h by a factor X, for example 2,048, and the output h/X is presented to a gate 56. The output d' is applied over a conductor 57 to a counter 58 where another factor Y, for example 4,096, is applied at the resulting output m,(d'/Y), is applied to the gate 56 to gate through the output l as a signal of h/X to the MCV display register of 32 which continues to receive, register and display the signal l until receipt of a stop signal from the predetermined count device 33, which occurs at, in this particular example, the count of 4096.

The counter 55 also applies a factor of Z to divide the signal h by, for example, 128 to provide the signal h/128 as a pulse series n to the total count volume display register 31 which is operable to receive, register and display counts during the start-stop interval initiated at the electrodes 20, 21 in FIG. 1. The counted signal h is therefore counted for the entire aliquot of sample and yields the factored HCT reading.

Although I have described my invention by reference to a specific application of a particular embodiment thereof, many changes and modifications of my invention may become apparent to those skilled in the art without departing from the spirit and scope of the invention, and it is to be understood that I intend to include within the patent warranted hereon all such changes and modifications as may reasonably and properly be included within the scope of my contribution to the art.

Claims

What I claim is:

1. In apparatus of the type wherein a liquid suspension of particles is caused to flow through a sensing zone, sensing means are provided to generate electrical pulses in response to the individual particles traversing the sensing zone for a predetermined interval, the particle pulses having amplitudes directly proportional to the volumes of the individual particles, means are provided for counting the particle pulses over the predetermined interval, and a data converter is utilized to register and display particle volume data in accordance with the amplitudes of the generated particle pulses, the improvement wherein said data converter comprises:

a pulse height analog to digital converter for converting the individual particle pulses into digital signals, means for summing said digital signals, means operable over the entire predetermined interval for applying a first factor to the summation of said digital signals to obtain total relative particle volume, and means operable over a predetermined number of particle pulses to apply a second factor to the summation of said digital signals to obtain mean particle volume.

2. The improvements set forth in claim 1, wherein said means for summing said digital signals includes an accumulator connected to said analog to digital converter, and said means for applying a first factor and said means for applying a second factor each include respective selected output taps of said accumulator.

3. In apparatus of the type wherein a liquid suspension of particles is caused to flow through a sensing zone, sensing means are provided to generate electrical pulses in response to the individual particles traversing the sensing zone for a predetermined interval, the particle pulses having amplitudes directly proportional to the volumes of the individual particles, means are provided for counting the particle pulses over the predetermined interval, and a data converter is utilized to register and display particle volume data in accordance with the amplitudes of the generated particle pulses, the improvement wherein said data converter comprises:

a pulse height analog to digital converter for converting the individual particle pulses into digital signals, means for summing said digital signals, and means operable over the entire predetermined interval for applying a factor to the summation of said digital signals to obtain total relative particle volume.

4. The improvement set forth in claim 3, wherein said means for summing said digital signals includes an accumulator connected to said analog to digital converter, and said means for applying a factor includes a selected output tap of said accumulator.

5. In apparatus of the type wherein a liquid suspension of particles is caused to flow through a sensing zone, sensing means are provided to generate electrical pulses in response to the individual particles traversing the sensing zone for a predetermined interval, the particle pulses having amplitudes directly proportional to the volumes of the individual particles, means are provided for counting the particle pulses over the predetermined interval, and a data converter is utilized to register and display particle volume data in accordance with the amplitudes of the generated particle pulses, the improvement wherein said data converter comprises:

a pulse height analog to digital converter for converting the individual particle pulses into digital signals, means for summing said digital signals, and means operable over a predetermined number of particle pulses for applying a factor to the summation of said digital signals to obtain mean particle volume.

6. The improvement set forth in claim 5, wherein said means for summing said digital signals includes an accumulator connected to said analog to digital converter, and said means for applying a factor includes a selected output tap of said accumulator.