US3520793A

US3520793A - Electrophoretic separator

Info

Publication number: US3520793A
Application number: US671894A
Authority: US
Inventors: Alexander Kolin
Original assignee: US Department of Navy
Current assignee: US Department of Navy
Priority date: 1967-09-29
Filing date: 1967-09-29
Publication date: 1970-07-14
Anticipated expiration: 1987-07-14

Description

July'l4,'1970 A. KOLIN ELECTROPHORETIC SEPARATOR 2 Sheets-Sheet 1 IMPROVED COLUMN Filed Sept. 29, 1967 PRIOR ART COLUMN INVENTOR. ALEXANDER KOLI N ERVl/V F JOHNSTON ATTORNEY y 1970 A. KOLIN 3,520,793

ELECTROPHORETIC SEPARA'I'OR Filed Sept. 29, 1967 2 Sheets-Sheet United States Patent 3,520,793 ELECTROPHORETIC SEPARATOR Alexander Kolin, Los Angeles, Calif., assignor, by mesne assignments, to the United States of America as represented by the Secretary of the Navy Filed Sept. 29, 1967, Ser. No. 671,894 Int. Cl. B01k 5/00 US. Cl. 204-299 6 Claims ABSTRACT OF THE DISCLOSURE The description discloses an improvement for an electrophoretic separator of the type wherein particles dissolved or suspended in a fluid are separated by subjecting the fluid to a combined action of a longitudinal electric field traversed by a perpendicular magnetic field within a horizontally extending migration column. The migration column is an endless fluid belt bounded on the inside by a soft iron core which is spaced from a surrounding jacket which forms the outer boundary of the fluid belt. On opposite ends of the fluid belt are disposed buffer chamhers which are capable of supplying a buffer medium for carrying the particles which are to be separated. This separator, which is fully described in Proceedings of the National Academy of Sciences, vol. 46 at page 509, utilizes a circular migration column as seen in cross section. Thepresent improvement employs a noncircular migration column, shaped like the belt of a belt sander, which overcomes, particle sedimentation problems.

The invention described herein may be manufactured and used by or for the Government of the United States of America for governmental purposes without the payment of any royalties thereon or therefor.

The ions within a biological or chemical mixture have different ionic mobilities which enable the ions to be separated when they are moved through an electric field. The apparatus for achieving such a result is an electrophoretic separator which has potential use in separating chemical mixtures or biological mixtures such as blood. For instance, the separation of blood into its components could isolate abnormal cells, cell debris, or virus particles which would be pertinent in diagnosing ailments.

In my patent application entitled Fractionation Apparatus, Ser. No. 500,817 and filed Oct. 22, 1965, now Pat. No. 3,451,918, I describe an electrophoretic separator which has a horizontal migration column which is circular in vertical cross section. Disposed at opposite ends of the migration column are a pair of chambers for providing a buffer flow which will carry a fluid under test through the migration column. Upon applying radial magnetic and axial electric fields with respect to the migration column, the solution or suspension under test will be separated into its various components within the column. This separator is also described in vol. 46 of the Proceedings of the National Academy of Sciences at p. 509.

Because of the difference of densities between the particles of the fluid under test and the carrying buffer solution the particles will tend to sediment by gravity to the .inner and outer walls of the migration column. With a circular migration column, which is used in my previously described electrophoretic separator, this problem is quite acute, as illustrated in FIG. 1. In order to avoid sedimentation in my previous separator it is necessary to maintain a rapid rate of rotational flow of the buffer containing the particles under test. I found that the sedimentation problems can be overcome by extending the vertical dimension of the migration column, as illustrated in FIG. 2. With such an arrangement the particles spend most of 'ice their time in vertical paths, where gravity causes no sedimentation toward a wall, and only a small fraction of their cycle in semicircular top and bottom paths, where there is a tendency to sediment toward a wall.

An object of the present invention is to provide an im provement of an electrophoretic separator which is described in my patent application entitled Fractionation Apparatus, Ser. No. 500,817, filed Oct. 22, 1965, now Pat. No. 3,451,918.

A further object is to provide an improved electrophoretic separator wherein sedimentation of particles flowing through the migration column thereof is minimized.

Other objects, advantages and novel features of the invention will become apparent from the following detailed description of the invention when considered in conjunction with the accompanying drawings wherein:

FIG. 1 is a schematic illustration of a cross-section of the migration column of my previous electrophoretic separator;

FIG. 2 is a schematic illustration of a cross-section of my improved migration column;

FIG. 3 is a schematic illustration, partially in crosssection, of an electrophoretic separator which includes my improved column; and

FIG. 4 is a view taken along plane IVIV of FIG. 3.

Referring now to the drawings wherein like reference numerals designate like or similar parts throughout the several views, there is shown in FIG. 3 my improved electrophroetic separator 10 which is an improvement of an electrophoretic separator described in my previously mentioned patent application. The electrophoretic separator 10 has a migration column 12 which is bounded by an iron core 14 and a surrounding jacket 16. The configuration of this column 12, which is illustrated schematically in FIG. 2, provides improved results of the separator over the configuration of my previous migration column, which is schematically illustrated in FIG. 1. As illustrated in FIG. 3, the improved migration column 12 extends horizontally and, as illustrated in FIG. 2, the migration column in vertical section has a large vertical dimension in contrast to its transverse dimension.

A fluid under test is injected into the migration column 12 from injector 68 and is carried through it along helical paths in a buffer solution. This fluid and solution will be described in detail hereinafter. During transition within the migration column the fluid particles traverse a magnetic field which is perpendicular to the surface of an iron core 14 and are subjected to a horizontal electric field which is perpendicular to the magnetic field and which causes separation of the particles according to their ionic mobilities.

Because of the difference in densities between the fluid particles and the carrier solution there is a tendency of the particles to migrate by gravity toward the walls of the migration column. FIG. 1, which is the prior art column, illustrates the deviation of a fluid particle which is more dense than the buffer solution from a center path after the particle has been discharged into the migration column 18. It can be seen that after injection the more dense fluid particle will, by gravity, first deviate inwardly toward the iron core 22 and will then subsequently deviate, also by gravity effect, toward the inner wall of the jacket 24. This deviation will cause the fluid particles which are more or less dense than the buffer solution to sedimentate on the walls of the migration column 18 unless rotational flow of the buffer fluid is maintained at a high rate. My improved migration column, as illustrated in FIG. 2, has overcome this problem by employing a noncircular configuration which causes the fluid particles to spend a greater amount of their time in

vertical paths

26 and 28, where gravity has no deviation effect, in contrast to top and

bottom paths

30 and 32, where there is a horizontal component. The deviation of the particles in the top and

bottom path portions

30 and 32 is so slight that it is not shown in FIG. 2. As can be seen from FIG. 1, the entire length of the particle path has a horizontal component which subjects the particles to gravity deviation.

The preferred configuration of my improved migration column 12, as seen in cross section is generally rectangular with top and bottom rounded ends, as illustrated in FIG. 2. It is important that the

legs

26 and 28 of the particle path be oriented vertically. The rounded top and

bottom ends

30 and 32 are preferably semicircular, as illustrated. Very satisfactory results can be achieved by employing an iron core 14 with the following dimensions; a horizontal length of approximately 7.5 cm.; a height of approximately 10.6 cm.; a thickness of approximately 2.6 cm.; and a radius of curvature at its top and bottom ends of approximately 1.3 cm.; the first dimension being illustrated in FIG. 3, and the latter three dimensions being illustrated in FIG. 2. The jacket 16, which may be constructed of Lucite, may be evenly spaced approximately 1.5 mm. about the iron core 14 so as to provide the migration column 12 with a uniform width of the same dimension about its vertical section, as seen in FIG. 2. In order to provide proper centering of the iron core 14 within the jacket 16 small cylindrical spacers (not shown) 1.5 mm. in thickness may be placed around the rim of the core 14 between the core and the jacket 16. It is important that the iron core 14 be insulated against electrical contact with the surrounding buffer solution, which will be described hereinafter. This insulation may be provided by painting the core with a spray varnish or by molding the core 14 into a crust epoxy which is subsequently milled off to leave a layer of a few thousandths of an inch adhering to the core 14 over its entire surface. If desired, the epoxy surface may be painted in suitable colors so that particle streaks may be easily viewed through the Lucite jacket 16.

Disposed at opposite vertical ends of the iron core 14 and the migration column 12 are

buffer chambers

34 and 36. The

buffer chambers

34 and 36 communicate electrically and hydraulically with one another through the migration column 12. Located adjacent the outer ends of the

buffer chambers

34 and 36 are

electrode compart ments

38 and 40 which may be separated from the buffer compartments by dialyzing

membranes

42 and 44. The dialyzing membranes 42 and 44 hydraulically isolate the buffer compartments from the electrode compartments but enable electrical communication therebetween. The buffer compartments 34 and 36 may be generally rectangular, as seen in vertical cross section (not shown), and the electrode compartments 38 and 40 may be divided into two sections 40a and 40b, as illustrated for the electrode compartment 40 in FIG. 4. These electrode compartment sections may be joined by a conduit 46 for hydraulic communication of the buffer solution, and may have

buffer overflow conduits

48 and 50 at their top ends. The buffer compartments 34 and 36 and the electrode compartments 38 and 40 may be formed -by Lucite walls, as illustrated in FIG. 3. The top of the buffer compartments 34 and 36 may be open so as to receive a buffer solution.

The buffer solution, which may be contained in a Mariotte bottle 52, is distributed to the butler compartments 34 and 36 and the electrode compartments 38 and 40 through various tubes and conduits. A plurality of thin plastic tubes divided into

bundles

54 and 56 may be utilized for distributing the buffer solution to the buffer compartments 34 and 36. The apportionment of these tubes among the

compartments

34 and 36 determines the rate and direction of axial buffer flow in the migration column 12. In actual practice these

tubes

54 and 56 are submerged into the buffer solution within the buffer compartments 34 and 46. The flow of buffer solution into these compartments may be cut by a stopcock 58 which opens into a manifold 60 and may be adjusted by varying the height of the Mariotte bottle 52. The buffer solution may be distributed from the Mariotte bottle 52 to the electrode compartments 38 and 40 through a conduit 62 which has branches opening respectively into the bottoms of the electrode compartments. A stopcock 64 may be employed for controlling the rate of flow of the buffer solution into the electrode compartments. The buffer solution may be composed of a pH 10 buffer tablet in one liter of water or some other type of solution of suitable pH suitably diluted may be used.

If a greater amount of the buffer solution is distributed to the buffer compartment 34 than the buffer compartment 36, there is a buffer flow to the left through the migration of column 12, as can be visualized from FIG. 3. Into this buffer flow is injected the fluid containing particles to be separated. This test fluid may be disposed within a reservoir and may be ejected into the migration column 12 near its upstream end by an injector 68 which is in communication with the reservoir 66 through a conduit 70. Accordingly, the fluid under test will be carried to the left within the migration column 12 by the buffer solution, as can be visualized from FIG. 3. During this flow the fluid under test, as well as the buffer fluid, is subjected to combined magnetic and electrical fields. The magnetic field may be provided by two pairs of bar magnets, one

pair

72 and 74 being disposed at the right end of the iron core 14 and the

other pair

76 and 78 being disposed at the left end of the iron core. Accordingly, the iron core 14 is sandwiched between the pairs of magnets. The pairs of magnets, which may be rectangular bars, are disposed with their polarities in opposition with respect to one another. Accordingly, the north poles of the pair of

magnets

72 and 74 may be placed in opposition with the north poles of the other pair of

magnets

76 and 78. If desired the south poles could have been used for opposition purposes. The magnetic field generated corresponds to the radial magnetic field of the circular configuration shown in FIG. 1. It penetrates the migration column 12 at right angles to the surface of iron core 14. The axial electric field through the migration column 12 may be provided by pairs of electrodes 80 and 82, one pair of electrodes 82a and 82b being disposed respectively within subcompartments 40a and 40b of the electrode compartment 40, as illustrated in FIG. 4, and the other pair of electrodes 80 being similarly disposed within the electrode compartment 38. The pair of electrodes 80 may be negatively charged and the pair of electrodes 82 may be positively charged by a potential source (not shown).

The pairs of magnets and the electrodes subject the fluid under test as well as the buffer solution to crossed electric and magnetic fields. This results in a force which makes the fluid in the migration column 12 circulate in the manner of motion of an endless belt. As a result of this circulation and the axial streaming to the left, the fluid particles describe a helical path. If the fluid under test has particles of varying ionic mobilities the electric field will add a horizontal component to their velocity and will separate these particles as they migrate to the left within the migration column 12 along spiral paths of dilfering pitch. The fluid particles of differing ionic mobilities will separate into individual streaks. A particle of slow ionic mobility is illustrated along the solid path 84 of FIG. 3, and a particle of faster ionic mobility is illustrated along the dotted path 86. These paths are helices of different pitch. The various particles of the fluid under test are then collected by a bundle of tubes 88 which have one series of ends disposed within the migration column 12 along a parallel to the axis of the iron core 14, and the opposite series of ends extending from the migration column 12 for collection purposes by a series of test tubes 90.

In order to suppress thermal convection of the fluid within the migration column 12 I have found it desirable to minimize the hydraulic communication of the buffer solution between the buffer compartments 34 and 36, and the migration column 12. This may be accomplished by a pair of nylon shoe laces 92 and 94 which are fitted tightly about respective opposite ends of the iron core 14 between the buflfer compartments 34 and 36. In order to allow unimpeded replenishment of the fluid leaving via the collector tubes 88 a small opening 96 may be cut into the left shoe lace 94 just below the collector tube ends.

In order to eliminate undesirable heating of the separator it} a cooling system may be employed for the migration column 12. The iron core 14 may be hollow so that ice water from a reservoir 98 may be perfused therethrough by inlet and outlet conduits 160 and 102. Further, the jacket 16 about the iron core may be provided with a peripheral chamber 104 which receives ice water from the reservoir 98 via a conduit 1%. The return from the chamber 104 may be via the conduit 102. In order to dispel air from the hollow core 14 when it is first charged with ice Water a conduit 108 and air outlet valve 119 may be connected to the top end of the iron core 14.

It is now readily apparent that the present invention will enable a more effective and efficient use of my previously described electrophoretic separator. By extending the vertical dimension of the migration column 12 the n particles to be separated will describe paths which are essentially unaffected by gravity deviation. Accordingly, the improved separator will have a much higher resolution and will permit processing of test fluids of a higher concentration. Another advantage is that fewer turns of the helix are required per given length of path of the injected particles in the new apparatus than was needed in the original circular version.

Obviously many modifications and variations of the present invention are possible in the light of the above 2 teachings. It is therefore to be understood that within the scope of the appended claims the invention may be practiced otherwise than as specifically described.

I claim: 1. In an electrophoretic separation apparatus which is stabilized against thermal convection, the combination of:

a base structure; a first tubular member mounted on said base structure and defining an electrophoretic migration column; said electrophoretic column being generally rectangular in vertical cross-section with top and bottom rounded paths and elongated paths therebetween, said elongated paths extending vertically; a first housing defining a first buffer fluid chamber mounted on said base at one end of the said tubular 7 member;

a second housing defining a second buffer fluid chamber mounted on said base at the other end of said tubular member;

said first tubular member intercoupling said first and second buffer fluid chambers for the flow of fluid through said tubular member from said first chamber;

first and second electrodes respectively positioned in said first and second chambers for producing an electric field longitudinally of said column between said first and second chambers so as to cause a migration of particle components of a substance to be separated along said column;

magnetic means for producing a magnetic field in said column extending generally radially across said column and traversely to said electric field to exert tangential forces on said particle components migrating along said column to cause said particle com ponents to assume different spiral-like paths;

a source of butter fluid coupled to said first and second butter chambers and having means for continuously supplying buffer fluid to said chambers and maintaining an essentially constant level of bufler fluid in said chambers to compensate for extraction losses and to maintain a predetermined axial flow distribution in said column;

means for injecting into said first tubular member a substance to be electrophoretically Separated by the buffer fluid flow in said migration column; and

pick-off means positioned in said migration column to intersect said particle components migrating along said spiral paths.

2. In an electrophoretic separator as claimed in claim It wherein:

each of the top and bottom rounded paths is semi circular.

3. In an electrophoretic separator as claimed in claim 2 wherein:

the dimension of the height of the electrophoretic column between its top and bottom semicircular paths is larger than the transverse dimension of width as measured between the centers of the two opposite vertical paths of the column.

4. In an electrophoretic separator as claimed in claim 2 wherein:

the dimension of the height of the electrophoretic column between its top and bottom semicircular paths is approximately four times the dimension of the width of the column between its vertical elongated paths.

5. In an electrophoretic separator as claimed in claim 4 wherein:

said electrophoretic column is formed by an iron core which is surrounded by a jacket; and including:

a thin layer of insulation bonded to said iron core.

6. In an electrophoretic separator as claimed in claim 5 wherein:

the iron core and jacket are both hollow; and including: means for circulating a coolant through the iron core and the jacket.

References Cited UNITED STATES PATENTS 3,207,684 9/1965 Dotts 204-480 3,287,244 11/1966 Mel 204-18O 3,305,471 2/1967 Miinchhausen et al. 204299 3,320,148 5/1967 Skeggs 204180 3,451,918 6/1969 Kolin 204299 JOHN H. MACK, Primary Examiner 0 A. C. PRESCOTT, Assistant Examiner US. Cl. X.R. 204180