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Патент USA US2412602

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Dec. 17; 1946.
. 1.. A. CHAMBERS EI‘AL
2,412,602
ELECTROPHORES IS APPARATUS
Filed Sept. 3, 1942
Qaafw guwiaow
2,412,602
Patented Dec. 17, 1946
UNITED STATES ‘PATENT oi-fncs)
2,412,602
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ELECTROPHORESIS APPARATUS’
Leslie Addison Chambers, Drexel Hill, and Haldan
Kelfer Hartline, Media, Pa., asslgnors to ‘The
Trustees of the University of Pennsylvania,
Philadelphia, Pa., a corporation of Pennsyl
vania
Application September 3, 1942, Serial No.v457,232
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7 Claims.
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(Cl. 88-14)
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often found in the form wherein the concentra
Our invention relates to optical interferom
eters, and particularly to their adaptation to the
analysis of the kinds and concentrations, relative
tion gradients are produced by a ?xed voltage
gradient (electrophoresis), it is not our intention
to in any way limit our invention to this embodi
ment, which we use below solely to illustrate our
or absolute, of particles in suspension or solution
in a liquid medium. Analysis is accomplished
through interferometric measurement of local
invention.
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Referring to the accompanying drawing, Fig
changesvin the refractive index of the suspension
or solution produced by the imposition of electri
cal, centrifugal or gravitational ?elds, or by dif
ure 1 is a side elevation of one possible arrange
ment of the interferometer unit for the applica
fusion. Any suspension or solution which can be 10 tion of a ?xed voltage gradient; Fig. 2 is a typical ,
schematic diagram of the essential components of
caused by any of the above means to exhibit such
the optical system; Fig. 3 illustrates the essential
local changes in refractive index may be analyzed _
‘light paths in the interferometer cell itself; Fig.
according to the method of our invention. .These
4 shows‘ the appearance of a typical photograph
local changes in refractive index are evidence ofv
concentration gradients.
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of the projected image of the interference fringes
.
In one of the methods of the practice of our
showing a concentration gradient between por
invention, concentration gradients are obtained
by permitting the migration of charged particles
of solute through a ?xed voltage gradient. The
several kinds of particles move in direction and 20
with velocity determined by the nature of their
tions of the solution being studied, the photograph
charge. This usually results in a stratification of -
the various kinds of solute particles. (If concen
tration gradients are produced by centrifugal or
gravitational ?elds, the rate and extent of strati
?cation are determined by the relative masses of
the solute particles.) , The formation, rate ~01
being taken from the front of one of the cells;
and Fig. 5 is a section taken on line 5—5 of the
cell shownin Fig. 1.
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Measurement of the interference fringe spacing
and displacement is correlated with other infor
mation to disclose the mass, charge, size and con
centrations of the solutes involved.
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The essential features of the illustrated em
bodiment of_.our invention may best be ‘under
stood by referring to Fig. 2, wherein I represents
the interferometer-electrophoresis cell submerged
movement. number and magnitude of the local
in the temperature controlled bath 2. The inter
refractive-index gradients which are caused by
the concentration gradients are detected by the 30 ferometer cell is, illuminated and observed by the
optical system shown. Light from the lamp 3
interferometric method (to be described). Infor
(preferably a vapor lamp) passes through the
mation may thereby be obtained concerning the
?lter l to render it monochromatic, thence
mass, charge, size and/ or absolute or relative con
through the condensing lens 5 whichibrings the
centrations of the several solute constituents.
The interferometric method of‘detection com 35 rays to convergence in a small‘ spot on the reflect
ing surface-of'prism 8. The light is directed by
bines sensitivity with compactness.
the prim 6 to collimating' lens ‘I, which is located
One of the objects of ‘our invention is to provide
at-its focal distance from the point of ‘conver
an apparatus which a?ords greatly increased
gence. The collimated beam then falls‘ upon an
sensitivity and a large reduction in the size of
interferometer portion of cell i placed at approxi
the necessary apparatus over other apparatus and
mately twice the focal distance from lens 1. The‘
methods usually employed.
re?ected beam-is directed through aperture‘! and
Still another object of our invent-ion is to pro
comes to focus on the photographic plat-e8, which
vide an apparatus which will combine increased
sensitivity and reduced size with means for actu
is at approximately'twice the focal distance'from
ally‘separati'ng and isolating the various ‘solutes 45 lens ‘I and is thus're'ceiving a real image of cell I.
Alternatively, the cell I may be placed ‘closer to
in complex mixtures, for further study outside the
apparatus.
I
Another object of our invention is to provide an
apparatus for the analysis of serum and of such
complex body ?uids-as plasma, s‘ynovial ?uids,
pleural exudates', urine, etc., as an adjunct ‘in the
diagnosis of certain diseases, which combines
sensitivity and‘ compactness through the use of
the interferometric method of'measurement.
_ lens "I and aphotographic-objective- lens placed‘
at 8 to'form- a r'ealimage of the=interferénce
fringes on the plate 8.-
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_In Fig. 1 is shown one possible combination
of interferometer and electrophoresis ‘cell: The
interferometer portions [0, shown in ‘section as 1 Fig; 5, constitute the center sectionsof 'a‘closed
U tube of rectangular cross-section designated ‘
9,419,009
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the interferometer portions I 0. The cells com
prise transparent plates "-23 separated by
spacer members 21-28. Part II, made up of
glass plates, forms the bottom of the U and the
assembly l2 makes up the top of the ‘U. This
section has two outlet tubes l2. and I4, which are
used to communicate liquid in the U tube-under,
examination with two appropriate glass vessels
containing the electrodes for introduction of the
‘electric current. The joints between tubes l3
and I4 and the tubes on the electrode vessels are
usually sealed by means of pieces of rubber tub
ing. It will be noted that the entire assembly is
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other about a vertical axis, thus enclosing a col
umn of liquid having a wedge shaped cross-sec
tion. The actual dimensional difference in thick
ness from side 21 to side 28 is such as to give from
'10 to 20 fringes (5 to 10 wavelengths) across the
width of the cell with the solvent in the cell. In
order to assure fringes which are straight and
‘ parallel, the inner faces of plates [8 and 22 are
made ?at within a very small fraction of a light
wave (e. g., 1/20). The fringes are held parallel
to the sides of the cell by accurate control of the .
dimensions of the spacer members 21 and 28 be
tween the plates. The amount of the inclination
of surfaces 20-22 is so slight that this does not
in the familiar‘ form, in general, whereby the
center portions It may be displaced laterally 15 show in the drawing. However, if lines were
drawn through the planes of the surfaces 20-22
with respect to either or both of the-bottom and
these lines would intersect in a point substantially
top sections. In this way it is possible to isolate
remote from the cell. Fig. 4 is an illustration of
portions of the solution in the U tube whenever
the appearance of the interference fringes at the
’
The construction of the interferometer por 20 boundary of two strata wherein exists a con
tions of the cellis best considered by reference - centration gradient as is impressed on the photo
graphic plate 9.
to Fig. 3. This ?gure shows the construction of
We have found the method of our invention
what is known as the-“re?ecting"'or “autocolli
highly practical and efficient in. operation. We
mating" type of interferometer, and is used mere
ly to illustrate the type of interferometer, and 25 realize that modifications may be made in carry
ing out the method of our invention and we in
not to limit (the interferometer to this particular
tend no limitations upon our invention other
design. The apparatus can incorporate the
than may be imposed by the scope of the ap
"transmission" interferometer as well. The lat
pended claims.
.
ter type is so constructed that the light source
and photographic apparatus are on opposite sides 30 What we claim as new and desire to secure by
of the interferometer. It is not intended to limit - Letters Patent of the United States is as follows:
1. The method of analyzing solutions which
detection of the interference fringes to a photo
comprises con?ning the solution under observa
graphic surface. Any suitable means may be
tion between a pair of interferometer plates, sub
employed, such as a telescope, scanning photo
jecting the solution therebetween to electrophore
- electric cell, etc. Returning to Fig. 3, number
sis, passing rays of monochromatic light through
17 indicates a ray of light entering the interfer
~the solution, and recording the interference pat
ometer from lens ‘I. This ray passes through the
tern produced by said interferometer plates for
transparent plate I! and strikes a semi-re?ecting
determining the characteristics of the solution.
coating I 9 on inner surface 20 of Plate l8. This
coating I9 is of such thickness that it transmits 40 2. Apparatus for analyzing solutions compris
ing a source of monochromatic light, a cell for
part and re?ects part of the incident ray I ‘I in
the solution having plane wall portions, one of
a de?nite ratio determined by the theoretical
which extends in a plane that is angularly dis
analysis of the system. The transmitted portion
posed with respect to the plane of the adjacent
2| of ray l'l passes through the solution being
examined and strikes the totally re?ecting coat 45 wall portion to provide a con?ning cell for the so
lution having slightly diverging walls, means for
ing 22 on the inner surface of plate 23. The ray
directing monochromatic light through said last
is re?ected by this surface through the solution
mentioned wall for re?ection by each of said
once more and impinges on the partial mirror
walls, optical means for directing the re?ected
ll, where part of it is re?ected and part trans
mitted to form ray 2|. The re?ected portion of 60 light to a light sensitive photographic plate, and
means for effecting a migration of the particles
_ the ray suffers a number of multiple re?ections
in said cell for recording interference fringes es
between the two mirror surfaces. Each time it
tablished in said cell according to the character
strikes surface l9, however, it loses part of its
istics of the solution therein.
energy as a returning ray. These rays help to
increase (or decrease) the fringe intensity and 55 3. The method of analyzing solutions which
comprises con?ning the solution under observa
thus contribute to the contrast ratio or resolving
tion between a pair of interferometer plates, sub
power of the interferometer. There is a theo
.iecting the solution therebetween to electrophore
retical optimum ratio of re?ectance to transmit
sis, passing rays of monochromatic light through
tance of ?lm is at which value the interference
fringes are the brightest and sharpest. The por 60 the solution, and optically observing the inter
ference pattern produced by said‘ interferometer
tion of the original ray H which is re?ected
plates for determining the characteristics of the
from coating is is designated by 25. ‘With the
solution.
conditions of illmnination as outlined herein, rays
it is so desired.
4. Apparatus for analyzing solutions compris
24 and 25 are in‘ a state and position to cause
ing a source of monochromatic light, a cell for the
interference with each other. Whether they ac
solution having plane wall portions, one of which
tually interfere or reinforce each other depends
extends in a plane that is angularly disposed with
entirely upon the path difference of rays 24 and
respect to the plane of the adjacent wall portion
25, and in effect depends only upon the time re
to provide a con?ning cell for the solution hav
quired for ray 2| to traverse the solution in both
directions. This time is controlled directly by the 70 ing slightly diverging walls, means for directing
monochromatic light through said last mentioned
index of refraction of the solution at that par
ticular spot.
In order to obtain interference fringes which
wall for re?ection by each of said walls. optical
means for directing the re?ected light to an op
tical observing means, and means for effecting a
will be useful for measurement and recording
purposes, surfaces 2|! and 22 are inclined to each 1g migration of the particles in said cell for optically
2,412,802
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observing interference fringes established in said
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a totally re?ecting surface on the interior face of
the other 01' said walls, a source of monochromatic
cell according to the characteristics of the solu
tion therein.
light directed through the semi-re?ecting surface
5. A system of analysis comprising a, cell for
of said cell upon the re?ecting surface thereof,
solution under examination having a horizontally 5 and means for observing the interference pattern
tapered chamber for the formation of optical in
resulting from the re?ection of the monochro
terference fringes, a source of monochromatic
matic light between the cell walls.
light directed upon said chamber, the surface of
7. A system of analysis comprising a cell for
the rear wall of said tapered chamber being pro
solution under examination, said cell comprising a
vided with a re?ector for said monochromatic 10 pair of plane walls disposed in diverging relation
light after the passage thereof through said cham
with respect to each other, a semi-re?ecting sur
ber, and means optically aligned with said cham
face on the interior face of one of said walls, a
ber in substantially the same transverse plane
totally re?ecting surface on the interior face of
through said light source and said tapered cham
the other wall, a source of monochromatic light di
ber for observing the interference pattern result 15 rected upon the totally re?ecting surface of said
ing from the re?ection of the monochromatic light
last mentioned cell wall through the semi-re
between the cell walls.
?ecting surface of the ?rst mentioned wall of said
6. A system of analysis comprising a cell for
cell, and means optically aligned with said source
solution under examination, said cell comprising a
of monochromatic light for recording the inter
pair of plane walls, one of said walls being dis 20 ference pattern resulting from the re?ection of
posed in a plane at a slight angle with respect to
the monochromatic light between the cell walls.
the plane of the opposite wall, a semi-re?ecting
LESLIE ADDISON CHAMBERS.
surface on the interior face of one of said walls,
HALDAN KEFFER. HARTLINE.
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