U.S. patent number 3,764,512 [Application Number 05/249,592] was granted by the patent office on 1973-10-09 for laser scanning electrophoresis instrument and system.
This patent grant is currently assigned to The Singer Company. Invention is credited to Ivan A. Greenwood, Jesse C. Kaufman.
United States Patent |
3,764,512 |
Greenwood , et al. |
October 9, 1973 |
LASER SCANNING ELECTROPHORESIS INSTRUMENT AND SYSTEM
Abstract
An improved instrument and system is provided for determining
the electrokinetic, or zeta potential of dispersed particles in an
aqueous solution. The system to be described includes a helium-neon
laser, electrophoresis cell, a lens system for expanding and
focusing the laser beam to produce a focal spot within the cell, a
galvanometer driven mirror for reflecting the laser beam from the
lens system into the cell, electronic circuitry for applying a
potential across the electrodes of the cell causing the dispersed
particles to migrate from one electrode to the other electrode,
electronic scanning circuitry for the galvanometer driven mirror
and a viewing microscope. The operator controls the angular
displacement and rate of angular displacement of the mirror until
the focused laser spot, as viewed through the microscope, tracks
the particles in the cell. A control may be calibrated directly in
zeta potential. The determination of the zeta potential is useful
in determining the degree of stability of particle dispersion in
aqueous solutions, and the like. The function of the scanning
instrument and system of the present invention is to measure the
zeta potential of colloidal particles dispersed in a solution.
Inventors: |
Greenwood; Ivan A. (Stamford,
CT), Kaufman; Jesse C. (Yorktown Heights, NY) |
Assignee: |
The Singer Company (Little
Falls, NJ)
|
Family
ID: |
22944165 |
Appl.
No.: |
05/249,592 |
Filed: |
May 2, 1972 |
Current U.S.
Class: |
204/645 |
Current CPC
Class: |
G01N
27/44721 (20130101) |
Current International
Class: |
G01N
27/447 (20060101); B01k 005/00 () |
Field of
Search: |
;204/18R,299
;356/102,103 |
References Cited
[Referenced By]
U.S. Patent Documents
Primary Examiner: Mack; John H.
Assistant Examiner: Prescott; A. C.
Claims
What is claimed is:
1. An instrument for measuring the electrophoretic mobility of
particles dispersed in a liquid and under the influence of an
electric field, said instrument including:
an electrophoresis cell for receiving and holding a liquid sample
having electrophoretic responsive particles therein and in which
said particles migrate under the infludence of an electric
field;
means for directing an illuminating beam at said cell to illuminate
the particles migrating in the sample;
a microscope positioned to enable the particles in the cell as
illuminated by the beam to be viewed by an observer; and
means for scanning the beam to cause it to track the movement of
the particles in the cell.
2. The instrument defined in claim 1, in which said directing means
comprises a laser and a lens system for focusing the beam from the
laser into a focal spot within the electrophoresis cell.
3. The instrument defined in claim 1, in which said scanning means
comprises a mirror-galvanometer type beam deflector.
4. The instrument defined in claim 1, and which includes an
adjustable power supply connected to said electrophoresis cell for
producing an electric field within the cell.
5. The instrument defined in claim 3, and which includes electronic
circuitry coupled to said galvanometer for producing the scanning
action of the light beam, said electronic circuitry including
manual controls for controlling the angular displacements of the
beam and the rate of displacement.
6. The instrument defined in claim 2, in which said electrophoresis
cell has a rectangular cross section, and in which said lens system
focuses the light beam from the laser cell to a focal point in a
region of said cell in which the average electro-endosmotic
velocity is essentially zero.
7. The instrument defined in claim 5, and which includes modulating
circuitry in circuit with said electronic circuitry for introducing
an additional scanning action to the galvanometer so that the light
beam is moved to one side of its normal position for short periods
and to the other side of its normal position for periods longer
than said short periods.
8. The combination defined in claim 1, in which said scanning means
comprises a galvanometer, and which includes electronic circuitry
connected through said galvanometer for introducing a scanning
signal thereto, and an adjustable power supply connected to said
electronic circuitry and to said electrophoresis cell to introduce
a common adjustable voltage to said circuitry and to said cell.
9. The instrument defined in claim 8, and which includes a control
in said electronic circuitry for adjusting the scanning rate of the
beam in said cell.
10. The instrument defined in claim 9, in which said last-named
control is calibrated in zeta potential.
11. The instrument defined in claim 9, in which said last-named
control is calibrated in mobility.
Description
RELATED COPENDING APPLICATION
(K-1888) - Laser-Retical Electrophoresis Instrument, Robert A.
Flower and Ivan A. Greenwood, Jr.
BACKGROUND OF THE INVENTION
The general expression for the zeta (electrokinetic) potential
is:
= 4 .pi. (300) .sup.2 .eta. v/.epsilon. E
= 36 .pi. .times. 10.sup.4 .eta. v/.epsilon. E
where:
Symbol Parameter Units .zeta. Zeta potential volts .eta. viscosity
poise v transfer velocity cm/sec .epsilon. relative dielectric
constant -- E potential gradient volts/cm
For an electrophoresis cell of the type employed herein, in which a
potential, V (in volts) is applied across the electrodes having an
effective spacing, d (in cm), equation (1) can be rewritten:
.xi. = 36.pi..times.10.sup.4 .eta.vd/.epsilon. V (2)
or in terms of electrophoretic mobility, .mu. (in microns/sec per
volt/cm)
.xi. = 36.pi..mu. (.eta./.epsilon.) (3)
It is noted that the parameters, viscosity and relative dielectric
constant are both temperature dependent. Laboratory temperatures
generally vary between 20.degree. C (68.degree. F) and 25.degree. C
(77.degree. F), so that zeta potential calibration should be
performed at a known, constant temperature. Typically,
Relative Dielectric Temp., T Viscosity, .eta. Constant, .epsilon.
20.degree.C 1.0019 .times. 10.sup.-.sup.2 P 80.36 25.degree.C 0.892
.times. 10.sup.-.sup.2 P78.54
solving equation (3), the zeta potential (which is usually
expressed in millivolts) is:
.xi. = 14.1 .mu. mV at T = 20.degree.C (4) .xi. = 12.9 .mu. mV at T
= 25.degree.C (5)
wherein the mobility has been expressed in the aforementioned units
of microns/sec per volt/cm. Equations (4) or (5) may be found
useful in converting electrophoretic mobility into zeta potential
in convenient units.
The instrument calibration initially required determination of the
maximum anticipation transfer velocity to which the laser focal
spot rate of displacement (scanning velocity) was to be matched.
Solving equation (2) for v under the conditions (T = 20.degree.C) ,
.xi. = 100 mV, V = 400 V, d = 10.3 cm, .eta. = 1.0019 .times.
10.sup.-.sup.2 P and .epsilon. = 80.36, a maximum beam linear
velocity, v.sub.max, of
v.sub.max = .xi..epsilon.V/36.pi. .times.10.sup.4 .eta. d (6)
= 2.75 .times. 10.sup.-.sup.2 cm/sec
was determined. The distance from the beam deflecting mirror to the
center of the electrophoresis cell was .rho. = 17.3 cm, thus the
corresponding maximum angular velocity, .OMEGA..sub.max, of the
galvanometer mirror deflector was
.OMEGA..sub.max = v.sub.max /.rho. (7)
= 1.59.times.10.sup.-.sup.3 rad/sec
= 9.11.times.10.sup.-.sup.2 deg/sec. A sweep period of t = 11.2
seconds was employed, thus the maximum angular deflection,
.theta..sub.max, was
.theta..sub.max = .OMEGA..sub.max t (8)
= 1.02.degree.
The angular sensitivity, (.DELTA.I/.DELTA..theta.), of each of the
two coils of the galvanometer was measured over the range 0.degree.
.ltoreq. .theta. .ltoreq. 18.degree. and in particular over the
range .theta. .ltoreq. .theta. .ltoreq. 2.degree.. The angular
sensitivity, over the latter range was
.DELTA.I/.DELTA..theta. = 14.0 mA/deg.
Thus, the maximum change in drive current, .DELTA.I.sub.max,
was
.DELTA.I.sub.max = (.DELTA.I/.DELTA..theta.) (.theta..sub.max)
(9)
= 14.3 mA.
With the "HI" and "LO" calibration potentiometers (R116 and R117)
set in their median position, the zeta potential potentiometer
(R118-A) control set at maximum (.xi. = 100 mV) and the adjustable
high voltage set at maximum (+ 400 Vdc), the precision current sink
parameters were adjustable to provide a coil (G101A) drive current
change of 14.3 mA during the sweep period.
Since the sensitivity of the beam position potentiometer (R118-B)
control was to be three times that of the zeta potential
potentiometer (for "double-gripping" averaging purposes) a similar
procedure was employed for the precision current sink driving the
other coil (G101B), resulting in a drive current change of 3
.times. 14.3 mA = 42.9 mA.
The zeta potential is an indication of the electrophoretic mobility
of the particles, which, in turn, is an indication of the velocity
with which the particles pass through the solution under the effect
of an applied electric field. The system of the invention measures
the rate at which the particles pass through the solution under the
influence of an applied electric field, and by that measurement
provides an indication of the zeta potential.
The apparatus and system of the invention finds particular utility,
for example, in the water processing industry. For example, the
instrument of the invention is useful in the industrial processing
of colloidal suspensions, and particularly in the purification of
industrial and drinking water.
When an electric potential is applied to the electrodes of an
electrophoresis cell, the charged particles migrate under the
influence of the resulting electric field toward the electrode of
opposite polarity. This migration can be observed under a
microscope, and such a measuring technique is standard in the prior
art electrophoresis apparatus. Such apparatus is described, for
example, in U.S. Pat. No. 3,454,478, which issued July 8, 1969, in
the name of Thomas M. Riddick. The electrophoretic mobility, that
is the velocity of the particles per unit field strength, is
measured in the Riddick apparatus by timing the particles viewed
through the microscope across a fixed distance in an observation
chamber under an electric field of known strength. By this process,
the mobility of the individual particles of different shapes and
sizes may be measured, and the corresponding zeta potential may be
determined.
However, the prior art cell, and its associated measuring system,
has limited feabilility in determining zeta potential in the case
of finished quality water, in which the number of colloidally
suspended particles is relatively small. The small number of minute
particles cannot be adequately observed in the microscopic field of
this instrument unless the operator is extremely expert in setting
up and viewing the necessary dark field illumination and, in any
event, the instrument is difficult to use under all conditions.
Further, the prior art instrument requires a manual conversion from
the time measurement into zeta potential, with a different
conversion factor being required for different electric fields.
Another problem in using this instrument is that only those
particles close to a reticle line can be counted, and this
limitation severely restricts the availability of particles which
can be used with adequate accuracy. Moreover, the measurements
require a high degree of skill on the part of the operator. Other
electrophoresis systems existed prior to this instrument, and
these, for the most part required an extremely short depth-of-focus
microscope, and the setting of the proper focal plane of the
microscope in such prior art systems was a tedious operation and
subject to inaccuracies.
The migration of the colloidal particles in the aqueous solution
occurs because charged groups and ions absorbed on the surface
produce a non-uniform distribution of ions in solution at the
particle-liquid interface. This distribution is generally expressed
in terms of zeta potential (millivolts) which may be calculated
from the mobility of the particles, as described above.
The present invention provides an improved system and instrument,
whereby the electrophoretic zeta potential of the particles in the
electrophoresis cell is measured by tracking the particles by a
scanning laser beam, and by the appropriate calibration of the
controls which are adjusted so that the laser beam accurately scans
in synchronism with the particles.
The present invention provides a semi-automatic system and
apparatus for measuring zeta potential. The use of the
semi-automatic instrument of the present invention is advantageous
in that it serves to reduce human error, and thereby improves the
speed, accuracy and reliability of the measurements, as compared
with the time consuming prior art techniques, such as described in
the Riddick patent, in which visual observations, through a
microscope, of the particle movement had to be timed and
tabulated.
As described above, the elements of the instrument and system to be
described include a laser, an electrophoresis cell of square or
rectangular cross-section, a beam expander and focusing lens system
adjustable to produce a laser beam focal spot within the
electrophoresis cell, a mirror-galvanometer type beam deflector for
reflecting the laser beam into the cell, a viewing microscope, an
adjustable voltage supply to produce a known electric field within
the electrophoresis cell, and electronic circuitry for producing
linear displacements and rates of linear displacement of the laser
beam focal spot under the control of two manually operated
controls. One of the controls represents displacement, and the
other represents zeta potential. The electrophoresis cell used in a
constructed embodiment of the invention, by way of example, had a
square configuration with a 5 mm .times. 5 mm internal dimension
and with a 10.3 centimeter effective length.
As will be described in more detail subsequently, the operation of
the apparatus of the invention is as follows. The electrophoresis
cell is filled with the aqueous solution having colloidally
suspended particles whose zeta potential is to be measured.
Electrodes composed, for example, of platinum-iridium (cathode) and
molybdenum (anode), are placed in the cell in contact with the
solution.
The operator then observes through the microscope the slowly moving
focal region of the laser beam inside the electrophoresis cell, and
he adjusts the sweep rate of the beam in the focal region so that
it appears to move at the same rate as the particles. This may be
achieved by adjusting the scanning rate control until good tracking
is observed upon repetitive sweeps of the beam. This may be
effectuated more rapidly by adjustment of the displacement control
until the beam is centered on an individual particle at the start
of its sweep, and then by adjustment of the rate control until the
beam tracks the particle.
Yet another technique is to set the laser beam on a particular
particle at the start of each of its sweeps by means of the
displacement control, and then by subsequently tracking the
particle with a "double gripping" procedure by which both the rate
and displacement controls are actuated at the same time. The latter
technique permits averaging over a number of particles when the
procedure is repeated over several sweep periods. In any event, the
rate of sweep generated by the rate control is multiplied by a
factor proportional to the applied voltage so that the rate control
may be calibrated directly in zeta potential. Under normal
conditions the same zeta potential setting will be obtained
independently of the voltage applied, which is of considerable
convenience to the operator.
The system and apparatus of the present invention has certain
advantages over the prior art systems, previously mentioned. For
example, the use of laser illumination permits even an
inexperienced operator to observe the particles clearly even in
situations where the particles are not visible in the prior art
instrument, as is the case of high quality finished water.
Moreover, once the tracking dials in the system of the present
invention have been manipulated to synchronize the scanning laser
beam with the migration of the particles within the cell, zeta
potential may be read directly off the dial without any additional
manual conversions, as is the case with the prior system.
In the system of the present invention the voltage across the
electrophoresis cell may be adjusted without affecting the final
zeta potential calibrations on the rate control dial. This permits
the voltage across the cell to be set so that a balance may be
found between a conveniently large particle velocity, and the
absence of extraneous convective particle movement caused by heat
transfer to the liquid by the conductivity currents which flow.
That is, the voltage across the cell can be set for optimum cell
conditions, without affecting the actual zeta readings.
In a preferred form of the instrument of the invention, the
electrophoresis cell has a rectangular cross-section, and the laser
focal spot traverses the cell along its length direction in a
region where the average electro-endosmotic velocity of the
particles is zero. The electro-endosmotic velocity arises from the
fact that the liquid in the cell is set into motion by interaction
with the cell wall and the electric field, independent of its
particle content. Since after reaching equilibrium there can be no
net liquid flow within the cell, there results a flow in one
direction near each cell wall and a counter flow down the center of
the tube. A closed, curved surface parallel to the cell
cross-section centerline defines the region wherein the two flows
balance out to zero, that is, as mentioned, where the
electro-endosmotic velocity is zero. Then, particle velocity
observed in a region along the curved surface of zero
electro-endosmotic velocity and in the direction of the cell
length, may be attributed completely to migration of the particle
by electrophoresis, and not by any other extraneous cause.
In a preferred embodiment, therefore, the laser focal spot sweeps
across the electrophoresis cell, along the aforementioned curved
surface (representing zero electro-endosmotic velocity) near the
top of the cell midway between the front and rear cell walls.
Measurements are made by observing only those particles illuminated
by the laser focal spot.
In the embodiment of the invention to be described, a coded angle
modulation may be added to the basic scanning motion of the mirror
so that the laser beam is moved to the left of its normal position
for short periods and to the right of its normal position for
longer periods (or vice versa) with a partial overlap of the two
deflected positions. Therefore, a particle exactly centered in the
undeflected position of the beam shows superimposed dots and dashes
giving a continuous unblinking illumination. A particle on one side
of the beam would then blink in dots, and a particle on the other
side of the beam would blink in dashes. Such a technique is
advantageous in the case where isolated particles are to be
measured, for it permits a more precise setting of the beam to the
selected particle since the width of the non-blinking zone is
typically much smaller than the width of the beam itself. Another
control which was found to be helpful was that of a variable
aperture around the beam which enables the operator to control the
beam size and brightness.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1(a) is a plan view of an apparatus and system incorporating
the concepts of the invention;
FIG. 1(b) is an elevation view of an electrophoresis cell as used
in this invention.
FIG. 2 is a perspective representation of an electrophoresis cell
positioned to be controlled in accordance with the present
invention, and an associated microscope for observing the particles
within the cell;
FIG. 3 is a block diagram representing the concept of the invention
on a functional basis;
FIGS. 4 and 5 show in circuit detail appropriate electronic control
systems for the invention; and
FIG. 6 is a circuit diagram of an appropriate modulation for the
galvanometer mirror, for the purposes described above.
DETAILED DESCRIPTION OF THE ILLUSTRATED EMBODIMENT
The instrument and apparatus of the present invention as shown in
the schematic representation of FIG. 1(a) includes, for example, a
laser 10. The laser may, for example, be a Spectra-Physics Model
131 Helium-Neon Gas Laser which produces a laser beam with a
wavelength equal to 632.8 nM. Even in finished water, particles are
clearly visible under laser light at the aforesaid wavelength.
A lens system 12 is provided in conjunction with the laser 10 for
expanding and focusing the collimated beam 11 from the laser into a
focal spot 19 at the selected region in an electrophoresis cell 14.
As mentioned above, a variable aperture 15 may be provided in
conjunction with the focused beam 13 to enable the operator to
control the beam size and brightness. The laser beam is reflected
into the cell 14 by a mirror 16 which is controlled, for example,
by a galvanometer 18.
Referring to FIG. 1(b), the cell includes a passage 20 which
extends longitudinally through the base from a first sample
receiving cup 22 to a second sample receiving cup 24. Electrodes 26
and 28 are positioned in the respective cups in direct contact with
the test liquid in the cell. As mentioned above, the cathode
electrode may be platinum-iridium and the anode electrode may be
molybdenum or of other composition chosen to reduce the formation
of gas bubbles within the liquid volume.
The light beam from the laser 10 is deflected by the mirror 16 in a
plane 21 (FIG. 1(b)) within the cell 14 and is focused in a focal
plane 17 (FIG. 1(a)) also within the cell. The intersection of
planes 17 and 21 preferably occurs in the zero electro-endosmotic
region. A microscope 30 (FIG. 2) is provided, for example, which
may be focused at plane 21 by means of the focusing adjustment 27.
The cell is supported, for example, on a support 32 which may be
composed, for example, of aluminum, or other appropriate material.
The microscope 30, for example, may be a Bausch and Lomb
"STEREOZOOM" Binocular microscope, although a monocular microscope
is satisfactory.
As shown by the block diagram of FIG. 3, the laser 10 is under the
control of a primary power supply 40, and the electrodes 26 and 28
of the electrophoresis cell 14 are under the control of an
adjustable high voltage power supply 42. As indicated by the block
diagram of FIG. 3, the laser output at .lambda. = 632.8 nM is 1.5
mW in the TEM.sub.00 mode. The beam expander and focusing lens
assembly 12 is secured to the laser housing. A threaded barrel
provides focusing range adjustment. The mirror-galvanometer
assembly 16, 18 which forms the beam scanner is oriented so that
the reflected beam is approximately normal to the incident laser
beam as shown in FIG. 1(a). The laser beam is deflected in the
horizontal plane 21, tending a full angle of 1.0.degree. (maximum)
when the high voltage control of the adjustable high voltage power
supply 42 is set, for example, to +400 V dc, and when the zeta
potential control 44 is set to .+-. 100 mV dc, as explained
above.
The electrophoresis test cell 14 is oriented in the horizontal
plane with its long dimension approximately normal to the reflected
laser beam. The focal region of the laser beam is nominally midway
between the cell electrodes 26 and 28 of FIG. 1(A), and precisely
midway between the front 23 and rear 25 inside cell walls. In a
constructed embodiment, and as mentioned above, a square cell is
used having internal cross section dimensions of 5.0 mm .times. 5.0
mm, where the effective spacing between the electrodes 26 and 28 is
approximately 10.3 centimeters. The height of the cell is adjusted
so that the laser beam traverses the square cross-section of the
cell tangentially to its upper zero electro-endosmatic region
located at plane 21 (FIG. 1(b), which is approximately 0.3 mm below
the internal top wall 29, of the cell.
The microscope viewer 30 is placed directly above the laser focal
spot 19 within the cell 14 and is manually focused to include plane
21 in the cell. The microscope 30 is located so that the scanned
laser beam is within the field-of-view of the microscope when the
cell voltage and zeta potential controls are set to a maximum.
The electronic circuitry for the instrument is shown functionally
in FIG. 3 within the broken line enclosure. The circuit details of
the electronic system are shown in FIGS. 4, 5 and 6.
The line voltage (110/120 volts AC, 50/60 Hz), from primary power
supply 40 is converted to fixed direct current voltages of +5V dc
and .+-.15V dc by power supply module 46 and to an adjustable high
voltage of 0-400V dc by the module 42. The low voltage power
supplies 46 provide circuit power and fixed reference voltages,
whereas the high voltage power supply 42 provides adjustable
control voltages as well as the potential for the electrophoresis
test cell 14.
The power supply 42 supplies a control voltage for a precision
sawtooth generator 50. A sawtooth waveform is generated by the
sawtooth generator 50 having a sweep period, for example, of 11.2
seconds and a flyback period of several milliseconds. The amplitude
of the sawtooth waveform is proportional to the output voltage of
the adjustable power supply 42. Therefore, the slope of the
sawtooth ramp is always proportional to the voltage gradient across
the electrophoresis cell 14, since both the sawtooth generator 50
and the cell 14 are supplied from the same power supply 42.
The sawtooth waveform from the generator 50 is applied to the
precision zeta potential control represented by the block 44. A
fraction of the sawtooth waveform voltage is then applied to a
precision current sink, represented by the block 52, which drives
one of the two galvanometer coils in the beam scanner 16, 18.
The other galvanometer coil is driven by a precision current sink
54 whose input voltage is derived from a summing amplifier 56. An
input signal to the summing amplifier is derived from the fixed
voltage power supply 46, independent of sweep direction. A second
input signal to the summing amplifier 56, is also provided by the
power supply 46 through a segment of switch function 53 which
applies this second input for one beam sweep direction only. A
third input signal is applied to the summing amplifier from the
adjustable power supply 42 through an inverter 57, through a sweep
direction segment of switch function 53, and through a beam
position control 58. This third input signal may be positive, zero
or negative, and its amplitude will depend both on the setting of
the beam position control 58 and the setting of the adjustable high
voltage power supply 42.
In the operation of the system, the electrophoresis cell is cleaned
and filled with the sample solution and secured to the holder 32
under the microscope 30. The zeta potential control 44 is set to
any convenient setting, possibly mid-range and the beam position
control 58 is set to approximately mid-range. The high voltage
adjustment control of the power supply 42 is adjusted to some
suitable level. The laser 10 is then turned on and the microscope
focus 27 is adjusted for the clearest image of the laser
illuminated particles in the region of the scanning laser focal
spot 19. The relative rates of beam scanning and particle migration
may be observed. The zeta potential control 44 is then adjusted
until the beam scanning and the particle migration rates are the
same. If this cannot be accomplished, the sweep direction/polarity
switch function 53 must be actuated to reverse the sweep direction.
This causes the polarity of the zeta potential voltage readout to
be of reversed sign. As mentioned above, the zeta potential control
44 can be adjusted alone, or in conjunction with the beam position
control 58, or the "double gripping" procedure may be followed.
When suitable tracking has been effected, the zeta potential, in
millivolts, of the solution under test is read out directly on the
dial of the zeta potential control 44. The polarity of the zeta
potential is indicated by the position of the sweep
direction/polarity switch function 53.
Referring to FIG. 4, the line voltage is supplied to the power
supply circuits by means, for example, of a power plug 100. The
plug is connected through a fuse F101 and through a power switch
SW101 to the low voltage power supplies 46. As described above, the
low voltage power supplies provide +5 volts and .+-.15 volts dc.
The power switch is also connected to an auto transformer T101
which supplies an adjustable alternating voltage to the primary of
a power transformer T102.
The voltage appearing across the secondary of the transformer T102
is rectified by a pair of diodes D101 and D102. The resulting
pulsating DC voltage is filtered by the usual filter circuit
including a choke coil L101 and a pair of capacitors C101A and
C101B. The resulting direct current voltage appears across the
resistor R101, and is applied through a high voltage fuse F102 to
the high voltage output terminal designated HV.
The output is shunted by a voltmeter VM101, so that the adjustable
voltage level of the high voltage power supply may be displayed.
The output voltage is also applied to the test cell 14 through a
switch SW102B. The switch SW102B is mechanically coupled to a
switch SW102A, the latter switch being connected in circuit through
a neon pilot lamp 60, and resistor 62, across the transformer T101,
as shown. As explained above, the low voltage power supplies 46
provide circuit power and fixed reference voltages, whereas the
high voltage power supply 42 provides adjustable control voltages
for the test cell and certain elements of the electronic
circuit.
As shown in FIG. 5, the precision sawtooth generator 50, is formed
by a precision integrating circuit made up of a pair of operational
amplifiers Z101 and Z102, and a flyback circuit made up of voltage
comparator Z103 and one-shot multivibrator Z104, and the circuits
of transistors Q104 and Q105. The circuits are connected as shown,
and are interconnected through field effect transistors Q101, Q102
and Q103. The field effect transistors may be of the type
designated 2N4067, the amplifier Z101 may be of the type designated
.mu.A741C, the amplifier Z102 may be of the type designated LM308D,
the comparator Z103 may be of the type designated .mu.A710C, and
the integrated circuit Z104 may be of the type designated 9601,
manufactured by Texas Instrument Company and other semiconductor
manufacturers.
The amplitude of the sawtooth waveform generated by the sawtooth
generator is proportional to the output voltage of the adjustable
power supply 42, so that the slope of the sawtooth ramp is always
proportional to the voltage gradient across the electrophoresis
test cell 14, as is desired for the reasons explained above.
The switch SW103 permits the sawtooth generator 50 to be disabled
when desired. The full sawtooth waveform is applied to the
precision zeta potential control potentiometer R118A through two
calibration potentiometers R116 and R117. A fraction of the
sawtooth waveform voltage is applied to the precision current sink
52 which is made up of an operational amplifier Z105, a
field-effect transistor Q106, and an NPN transistor Q107, connected
as shown. The operational amplifier Z105 may be of the type
designated .mu.A741C, the transistor Q106 may be of the type
designated 2N3456, and the transistor Q107 may be of the type
desigated 2N2222. The resulting output from the precision current
sink 52 drives one of the coils G101A of the galvanometer 18
through a reversing switch SW104 included as part of switch
function 53.
The other galvanometer coil G101B is driven by the precision
current sink 54 which is made up of an operational amplifier Z108,
a field-effect transistor Q108 and an NPN transistor Q109, which
may be the same as corresponding elements in the precision current
sink 52. The input to the precision current sink 54 is derived from
the summing circuit 56 which is made up of an operational amplifier
Z107 which may be of the type designated .mu.A741C, and which is
connected as shown. The sweep direction/polarity switch function 53
further includes sections designated SW104d and SW104a in the
circuit of the operational amplifier 56, and in the connections of
the low voltage power supply 46 to the input of the summing
amplifier. The switch function 53 also has a further section
designated SW104b in the output of the inverter 57. The various
sections of switch function 53 are ganged together such that they
move in unison. The switch function 53, through its various
sections, enables the laser beam to be automatically swept either
to the right or to the left, provided that the automatic sweep
switch SW103 is closed and always within the field of view of
microscope 30.
As illustrated in FIG. 5, one input for the summing amplifier 56 is
derived from the +5 volt fixed voltage source 46 as applied through
a 240 kilo-ohm resistor R130, this input signal being independent
of the sweep direction. The second input signal to the summing
amplifier Z107 is supplied through the switch SW104A and through a
240 kilo-ohm resistor R133, and is provided only when the laser
beam sweeps to the left. The third input signal to the summing
amplifier 56 is derived from the adjustable high voltage supply 42
through the switch SW104b and through the beam position control
potentiometer R118B, and through a 240 kilo-ohm resistor R127.
The inverter 57, including Z106, which may be of the type
designated .mu.A741C, provides equal and opposite voltage to the
extreme positions of the beam position potentiometer R118B. The
beam position potentiometer R118B is mechanically coaxial to the
zeta control potentiometer R118A. The output voltage from the
potentiometer R118B is positive, zero or negative. The magnitude of
the voltage depends both on the beam position potentiometer setting
and the setting of the adjustable high voltage supply by the
autotransfromer T101.
As mentioned above, a coded angle modulation may be added to the
galvanometer scanning mirror 16 so that the laser beam may be moved
to the left of its normal position for short periods and to the
right of its normal position for longer periods, and this may be
achieved by interposing the modulator circuit of FIG. 6 between the
zeta potential control potentiometer and the input of the
operational amplifier Z105. When the switch SW105 of FIG. 6 is in
its upper position, the potentiometer R118A is directly connected
to the amplifier Z105, and the modulator of FIG. 6 is ineffective.
However, when the switch SW105 is in its lower position, the
operational amplifier Z112 is interposed between the potentiometer
R118a and the amplifier Z105.
The modulator of FIG. 6 is connected in the indicated manner, and
includes the operational amplifier Z112, and a second operational
amplifier Z111, each of which may be of the type designated
.mu.A741C. The modulator also includes a pair of one-shot
multivibrator circuits designated Z109 and Z110, each of which may
be of the type presently designated 9601. The circuit includes an
offset voltage adjustment potentiometer R142 which is connected,
together with resistor R141 and R143, across a direct voltage
source, of the order, for example, of 400 volts. The potentiometer
R142 is connected to the input of the operational amplifier Z111,
as is the output of the integrated circuit Z110. The modulator also
includes a pulse amplitude adjustment control in the form of a
potentiometer R148 at the output of the amplifier Z111, and which
is connected to the input of the amplifier Z112.
As mentioned above, the circuit of FIG. 6 operates to move the
laser beam to the left of its normal position for short periods and
to the right of its normal position for longer periods, with a
partial overlap of the two deflected positions. Therefore, when the
modulator is operational, a particle exactly centered in the normal
position of the beam shows superimposed dots and dashes giving a
continuous unblinking illumination. However, a particle on one side
of the normal position of the beam blinks in dots, and a particle
on the other side of the normal position of the beam blinks in
dashes.
The modulator is effective in the case of isolated particles in
assisting the adjustment of the system to a desired particle.
However, where the particle density is high, the displacement
modulation lights up other particles and distracts the operator, so
that the switch SW105 should be switched to its up position to
disable the modulator for such a situation.
When the modulation switch SW105 is moved to its lower or ON
position, a pulsed waveform is added in the amplifier Z112 to the
sawtooth waveform derived from the zeta potential potentiometer
R118a. The pulsed waveform is obtained from a free running
multivibrator formed by the interconnection of the circuits Z109
and Z110 as shown and having "on" and "off" periods, for example,
of approximately 0.1 second and 0.7 seconds, respectively.
The potentiometer R142, and the summing circuit formed by the
operational amplifier Z111 provide the desired zero voltage level
setting for the waveform. The second potentiometer R148 enables the
magnitude of the modulation to be optimized for a given dispersed
particle size and laser focal spot size combination.
The eventual result of the two waveforms applied to the amplifier
Z112 is a net motion of the mirror 16 consisting of a superposition
of two motions, namely, a slow, linear sweeping motion associated
with the sawtooth waveform and a relatively small scale, back and
forth motion associated with the coded angle modulation. When a
particle is located in the plane of the scan, but slightly off
center with respect to the normal beam focal position, the particle
is illuminated in a pulse fashion for periods of 0.1 seconds, if on
one side, and for periods of 0.7 seconds, if on the other side, the
illumination occurring every 0.8 seconds, insofar as a constructed
embodiment is concerned.
The blinking of the particle indicates to the examiner whether it
is leading or laggging the normal position of the scanning beam.
The change in modulation factor also indicates to the operator the
direction of the relative velocity between the particle migration
and the beam scan. Therefore, the operator may implement the
necessary corrective actions to cause the particles to remain
centered with respect to the normal beam position as indicated by a
continuous illumination of that particle, so that the desired
tracking may be effectuated for proper read-out of the zeta
potential.
The invention provides, therefore, an improved and relatively
simple system which enables an operator to determine the zeta
potential of dispersed particles. The system and instrument of the
invention is most advantageous in that it is capable of producing
accurate results, and yet does not require any high degree of skill
on the part of the operator.
Although the invention has been illustrated by describing a system
wherein the particle velocity is monitored in the plane of zero
electro-endosmatic velocity nearest the top of the cell, it is to
be remembered that the region of zero electro-endosmatic velocity
is in fact a closed curved surface, in close proximity to the cell
walls, which runs the length of the electrophoretic cell. Therefore
there are an infinite number of planes, which intersect this
surface, which could be monitored for purposes of this invention.
The only determinations for working with any of these other planes
would be the establishment of the geometrical relationships between
the reflected laser beam and the plane intended to be monitored and
between the microscope and said plane. For example, the plane
parallel to plane 21 of FIG. 1(b) and positioned approximately the
same distance (0.3 mm) above the lower cell wall would serve the
purposes of this invention provided the laser focal spot 19, was
positioned in that plane and microscope 30 was focused thereon.
Alternately, if the plane which is positioned midway between the
upper and lower cell wall is utilized and the laser beam and
microscope are positioned accordingly to focus in this plane, and
the microscope is provided with a suitably engraved reticle
defining the distance in from the cell wall corresponding to the
intersection of the closed curve defining the zero
electro-endosmatic velocity with the plane (as is done in the
instrument described in the Riddick patent) then the proper
measurements may be made by observing only those particles near the
reticle line. Although this requires additional operator judgment
than might be desired it does relax the requirement for laser beam
focusing.
Although in the preferred embodiment the laser beam is scanned at
the same rate that the particles move, with the cell and microscope
being fixed, it is to be appreciated that, alternatively, it is
possible to keep the laser beam fixed and to physically translate
the cell at a velocity equal to and oppositely directed from the
particle velocity. Further it is possible to spread the laser beam
(using a cylindrical lens, for example, interposed between the
reflecting mirror and the cell) into a broad but thin focal region
thereby illuminating a larger section of the zero velocity
electroendosmatic surface and to then track particle velocity by
means of a mechanically moved reticle in the microscope eye piece
or to use a scanned mirror in the optical path between the
microscope and the cell, with fixed reticle marks in the eye
piece.
It can also be appreciated that changes in the above embodiments
can be made without departing from the scope of the present
invention, and that other variations of the specific construction
disclosed above can be made by those skilled in the art without
departing from the invention as defined in the appended claims.
* * * * *