U.S. patent number 3,803,958 [Application Number 05/211,135] was granted by the patent office on 1974-04-16 for ultra thin sectioning with ultra sharp diamond edge at ultra low temperature.
Invention is credited to Humberto Fernandez-Moran.
United States Patent |
3,803,958 |
Fernandez-Moran |
April 16, 1974 |
ULTRA THIN SECTIONING WITH ULTRA SHARP DIAMOND EDGE AT ULTRA LOW
TEMPERATURE
Abstract
Ultra sharp diamond edge is used for molecular and submolecular
sectioning at ultra low temperature and as a high intensity point
source for the emission of electrons rons and neutrons.
Inventors: |
Fernandez-Moran; Humberto
(Chicago, IL) |
Family
ID: |
26905870 |
Appl.
No.: |
05/211,135 |
Filed: |
December 22, 1971 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
Issue Date |
|
|
829267 |
Jun 2, 1969 |
3646841 |
|
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|
466877 |
Jun 22, 1965 |
3447366 |
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Current U.S.
Class: |
83/15; 83/170;
83/915.5 |
Current CPC
Class: |
H01J
37/285 (20130101); G01N 1/06 (20130101); G01N
2001/068 (20130101); Y10T 83/283 (20150401); Y10T
83/041 (20150401); G01N 1/42 (20130101) |
Current International
Class: |
G01N
1/04 (20060101); H01J 37/26 (20060101); H01J
37/285 (20060101); G01N 1/06 (20060101); G01N
1/42 (20060101); B26d 007/08 (); G01n 001/06 () |
Field of
Search: |
;83/916.5,170,171,15 |
References Cited
[Referenced By]
U.S. Patent Documents
Primary Examiner: Meister; J. M.
Attorney, Agent or Firm: Burns; Robert E. Lobato; Emmanuel
J.
Parent Case Text
This application is a division of my application Ser. No. 829,267
filed June 2, 1969, now U.S. Pat. No. 3,646,841, which in turn was
a continuation-in-part of application Ser. No. 466,877, filed June
22, 1965, now U.S. Pat. No. 3,447,366.
Claims
What I claim is:
1. A method of making ultra-thin serial sections of a specimen,
comprising the steps of mounting said specimen on a first holder in
a chamber, mounting on a second holder in said chamber a diamond
knife having a cutting edge thickness of the order of 0.001 to 0.01
microns, drawing an ultra high vacuum in said chamber, cooling said
chamber, first and second holders, specimen and knife to a
temperature lower than 5.degree. K, moving said knife and specimen
relative in a first direction relative to one another to make a
first cut of said specimen with said knife, moving said knife and
specimen relatively toward one another in a second direction
transverse to said first direction a distance of 10A. to 100A.,
again moving said knife and specimen relative to one another in
said first direction to make a second cut of said specimen parallel
to the first cut, thereby cutting a section of said specimen having
a thickness of 10A. to 100A. corresponding to the relative movement
of said knife and specimen in said second direction, repeating said
movements alternately in said first and second directions to cut a
series of sections of said specimen and collecting said sections in
a bath of cryogenic fluid.
2. A method according to claim 1, in which said chamber, first and
second holders, specimen and knife are cooled with liquid helium
and in which said bath is liquid helium.
3. A method according to claim 1, in which one of said holders
comprises a thermally expansive member, and in which light is
directed on said member to cause expansion thereof to effect
relative movement of said knife and specimen in said second
direction.
4. A method according to claim 1, in which one of said holders
comprises a magnetostrictive means, and in which a magnetic effect
is applied to said magnetostrictive means to effect relative
movement of said knife and specimen in said second direction.
5. A method according to claim 1, comprising shielding said
holders, specimen and knife from radiation effecting temperature
changes.
6. A method according to claim 1, comprising cooling said holders
by thermal conduction through flexible thermal conductors.
7. A method according to claim 1, in which said specimen comprises
macromolecules, comprising cutting said macromolecules into
constituent components.
8. A method according to claim 1, further comprising isolating said
first and second holders from external vibration.
Description
It is widely recognized that ultra sharp diamond knives and
associated ultramicrotomy apparatus of the type developed and
systematically introduced in research laboratories and in industry
by the applicant have considerably advanced the state of prior art
in the preparation of very thin slices of materials, and in
ultraprecision machining. His pioneering work in this specialized
field has resulted in the production of the highest quality metal
surfaces presently attainable with finishes and tolerances in the
low-microinch range.
This unprecedented, readily attainable microinch accuracy is now
achieved directly by continuous turning operation using the diamond
knife, without the traditional time-consuming and very costly
additional work of lapping, buffing or polishing.
The cutting-edge sharpness radius of the order of 0.10 microinch,
and the regularity of the diamond knife cutting edge are of
decisive importance for this purpose, as compared with the measured
cutting-edge radius of well-sharpened conventional tools which is
about 300 microinches. As shown by careful studies, this far
greater cutting-edge radius and irregularity of the best standard
tools results in the production of high frictional forces on the
cutting edge which generates a poor surface finish by tearing and
deforming the material plus accelerated wear and abrasion of the
edge.
However, with a diamond cutting tool having an edge thickness of
0.001 to 0.01 micron results are consistently obtained that
practically approach the idealized condition for optimum
sectioning. Since the zone of compressive yielding and flow around
the edge is reduced to a vertical length of a few microinches, more
uniform flow and yielding of the material at the active tool edge
is achieved with consequent improvement in the surface finish of
the cut. Moreover, very precise sizing can be maintained because
cut depths of only 5 to 10 microinches are readily made.
It is therefore essential to determine accurately the dimensions
and properties of these ultra sharp cutting edges as an
indispensable factor in the production and testing of the diamond
knife, considered to be the most sophisticated cutting tool
available today.
Prior to my work in this field, there were no adequate methods for
direct imaging and measuring the cutting-edge sharpness radius of
dimensions below the resolving power of the light microscope (0.2
to 0.4 micron). In fact, the active cutting region of our sharpest
cutting tools remained "invisible" until suitable methods were
devised which permitted successful application of the hundredfold
higher resolving power of the electron microscope for direct
observation of these submicroscopic structures. Direct
visualization and precise measurement of the ultra sharp cutting
edges was an integral part of research program in ultramicrotomy,
and played a key role in the development of the diamond knife. The
critical measurement and testing procedures for the cutting edges
which have been developed in the course of this work represent the
basis for a rational approach to the complex problems encountered
in the new domain of ultraprecision machining and molecular
sectioning.
Conversely, the possibility of working with stable cutting edges of
submicroscopic dimensions, and of studying their interaction with
selected specimens under controled conditions has opened up a new
field of investigation, disclosing new phenomena which are
operative predominantly at the level of molecular organization.
Thus, for example, with a diamond cutting-edge sharpness radius
measuring only 10 to 100 A., which corresponds to some six to 60
carbon atoms strongly covalently bonded, it is actually possible to
section or "cut up" certain long-chain organic polymers into their
constituent chemical subunits of molecular dimensions. By using the
special ultramicrotome apparatus described earlier operating at
very low temperatures it has been possible to cut starch
macromolecules into their constituent sugar molecules endowed with
quite different chemical properties.
This controlled modification of molecular structures represents a
new form of manipulation, extending well beyond the established
definition of mechanical sectioning, and leading into the realm of
a novel modality of "preparative chemistry and physical chemistry"
performed under conditions of mimimum perturbation in a low entropy
environment.
The methods described in this application were used primarily in
connection with the production and testing of diamond knives; since
diamond is the hardest naturally occurring material, and its
crystalline structure makes it possible to reproducibly obtain
stable cutting edges of molecular dimensions, using appropriate
techniques and apparatus, of the type described in my U.S. Pat.
Nos. 2,961,908, 3,060,781, and 3,447,366.
However, the measurement and testing procedures can likewise be
applied to determine the sharpness and properties of the active
cutting edges of all kinds of cutting tools, including glass knives
for ultramicrotomy, razor blades and related steel cutting edges,
the wide variety of well-sharpened machine tools, and in general
all cutting tools used in science, industry and technology where
optimum sharpness and regularity are essential. It should be
pointed out that accurate determination of cutting-edge sharpness
radius and related properties of active cutting tools is of
fundamental significance as an integral part of any manufacturing
or production procedure designed to systematically achieve
efficient cutting tools for sectioning all types of materials, in a
controlled and economic way. This difficult field has been
neglected in the past, partly owing to technical limitations
inherent in light microscopy. However, now that ultraprecision
machining and related disciplines are assuming vital importance in
an era of increased automation, which depends for its success on
absolute uniformity of operation and reproducibility of the
finished product, the refined measurement techniques described here
are of key operational value. In many ways a similar relationship
can be invoked between the improvements in measuring techniques and
the subsequent spectacular advances in ultrahigh vacuum technology.
It was only after reliable devices and techniques for measuring
ultra-high vacuum, like the Alpert gauge, became available that the
whole field of attaining and monitoring ultrahigh vacuum
environments came into being and could be systematically developed
on a rational basis.
The invention relates generally to measurement and testing
procedures for ultra sharp cutting edges. The separate steps can be
carried out individually in different stages, but are best applied
sequentially in systematic combination, since they are mutually
supplementary, and finally yield an integrated picture of the
cutting edge configuration and properties.
The procedure embodies the following distinguishing features
incorporated in the successive operational stages:
a. Direct examination of the ultra sharp cutting edge and its
adjacent components, carried out step by step continuously at the
different levels of magnification extending from the range of light
microscopy to electron microscopy, electron diffraction and related
electron optical techniques, in a non-destructive, highly accurate,
and reproducible quantitative analytical way;
b. Supplementary examination of the surface characteristics of the
cutting edge facettes and relationship to the active cutting radius
of electron optical examination of a special type of high
resolution indentation replica especially developed for this
purpose;
c. Ultimate critical test of the performance and stability of
ultrasharp cutting edges by actual cutting of ultrathin serial
sections of selected specimens, such as metals, hard crystalline
substances, macromolecular materials and biological specimens with
a highly ordered internal structure. The quality and properties of
the cutting edge can be accurately evaluated by combined electron
microscopy, electron diffraction, physical, and physical-chemical
studies of the resulting ultrathin sections and the exposed
speciment block surface;
d. In special cases, and particularly for routine monitoring and
inspection of large numbers of cutting tools with ultra sharp
edges, certain procedures for examinations based on the
characteristic electron optical, electrical, and ion-optical
properties of these cutting edges can be adapted for automated
inspection on an assembly line basis.
In the following description of the apparatus end method in
accordance with my invention, reference is made to the accompanying
drawings in which:
FIG. 1 is a schematic perspective illustration of a diamond tip
point cathode source;
FIGS. 1A, 1B and 1C are enlarged schematic views of the diamond tip
point cathode source and associated parts of FIG. 1;
FIG. 2 is a characteristic brightness curve of the point source
illustrated in FIG. 1;
FIG. 3 is a schematic sectional view of a high intensity
field-emission electron and ion sources using an ultra sharp
diamond edge; and
FIGS. 3A and 3B are enlarged schematic views of the ion source
shown in FIG. 3;
FIG. 4 is a schematic perspective view of apparatus for molecular
and submolecular sectioning.
FIG. 4A is an exploded schematic view of a cryogenic motor of the
apparatus shown in FIG. 4.
a. Direct examination of the ultra sharp cutting-edge by high
resolution light microscopy and electron microscopy, and related
electron-optical techniques.
This procedure can best be illustrated by referring to a typical
example such as a diamond knife of the type described in my U.S.
Pat. No. 3,060,781. A representative diamond knife comprises a
diamond body portion with a perfectly uniform and stable cutting
edge of about 3 to 7 mm length, and with a cutting-edge sharpness
radius of about 10 to 100 A. units (0.001 to 0.01 micron). These
diamond knives usually have facettes defining edge angles of about
40.degree. to 50.degree. for cutting plastic materials, and of
75.degree. to 80.degree. for cutting metals and other hard
substances. However, contrary to certain claims, these edge angles
do not define the active cutting-edge of the knife. Instead, the
active cutting region is several hundred Angstrom units from the
edge line, since it is built up of the regularly disposed
crystalline unit layers of the diamond arranged in stable
configuration to give smooth facets. In order to carry out the
first series of optical tests the diamond knife embedded in its
metal support must be mounted in a specially designed holder which
permits it to be very accurately positioned and oriented at
preselected optimum angles for direct examination by light
microscopy, and subsequently by electron microscopy. A
photomicrograph of the edge taken at the highest light microscope
magnifications (1,000x to 2,000x) shows a perfectly regular,
straight and extremely smooth cutting edge facet. The actual
cutting edge is not directly visible, since it lies beyond the
resolving power of the ordinary light microscope. However, when
examined under optimum conditions with point light sources of very
high intensity (e.g., Xenon lamps, lasers, etc.) with the dark
field "ultramicroscope" the active cutting edge can be faintly
detected by virtue of its light-scattering properties. This allows
detection of small cracks and irregularities in the edge, but still
does not permit direct imaging of the critical active cutting edge.
Nevertheless, improved light microscopy techniques such as
interference microscopy permit accurate determination of the
general quality of the cutting edge and adjoining facet. This is
especially useful when comparing the uniform contours and regular
spacing of the drak interference bands indicating a perfectly plane
and smooth diamond edge and facet, with the curving and irregularly
distorted interference fringes corresponding to the submicroscopic
irregularities of a polished steel razor edge. Following these
initial observations which can be readily carried out routinely
during the various steps of the final polishing of the diamond or
other type of ultra-sharp cutting edge, the decisive electron
optical examination is carried out.
For this purpose the same holder can be used, suitably adapted for
introduction into the high vacuum specimen chamber of an electron
microscope. These studies require the use of a specially modified
high resolution electron microscope operating at accelerating
voltages of 40 to 100 kV in most cases (also in the low voltage
range of 5 to 10 kV for certain applications), provided with a
special pointed filament source with single-crystal tungsten tips
of the type developed by the applicant. By means of these pointed
sources highly coherent microbeam illumination (beam diameters
ranging from 100 A. to 10 microns) of low energy spread and very
low intensity can be used, in conjunction with suitable high or
ultrahigh vacuum specimen chambers preferably cooled with liquid
nitrogen or hydrogen (vacuum of the order of 10.sup..sup.-6 to
10.sup..sup.-8 mm Hg) to prevent the deleterious contamination of
the knife edge during illumination with the electron beam. Under
these optimum conditions the properly oriented cutting edge can be
very clearly seen directly and photographed at electron optical
magnifications ranging from 2,000x to about 200,000x, with
subsequent photographic enlargement to attain total magnifications
of half a million to about one million diameters. With a focused
electron microbeam of adequately low intensity to avoid specimen
irradiation damage the electron microscope can readily be employed
to produce a distinct shadow image of the diamond knife edge and
supporting region, at magnifications adequate to resolve fine
details of the edge in order to a few hundred Angstroms. Imaging by
reflection electron microscopy can also be readily accomplished by
using oblique illumination and tilting the knife so that the
incident electron beam makes a very small angle with its surface.
This simple technique yields valuable data on the surface structure
of the knife facet and of the active cutting edge, supplementing
the results obtained by examination of the surface replicas.
However, the most important information is obtained by
straightforward transmission electron microscopy with microbeam
illumination at high magnifications, essentially because the active
ultra sharp cutting-edge is ultrathin, and therefore it alone is
electron transparent and can be selectively imaged. This remarkable
effect which was discovered in the course of these studies has
provided to be of extraordinary value as one of the most direct and
powerful means of studying ultra sharp cutting edges at the highest
electron-optical magnifications. It is readily understandable,
since the ultra sharp cutting edges with a radius of merely 10 to
100 A are built up of a relatively small number of the crystalline
unit layers of diamond to form an ultrathin wedge-shaped structure
which is at most only a few hundred Angstrom units thick at the
base, and therefore stands out selectively as an "electron
transparent window" against the rest of the knife base which is
electron-opaque. The fact that the ultrathin active cutting-edge of
an ultra sharp cutting tool is essentially an "electron-transparent
window" is of unique operational value because it permits us to
study directly all structural details clearly visible in this most
important region under excellent conditions which minimize specimen
radiation damage. This effect is also clearly discernible when
using electron diffraction techniques, since there is a
well-defined transition from a reflection-type electron diffraction
pattern to a "transmission electron diffraction pattern" precisely
along the boundaries of the ultrathin cutting-edge. Although these
characteristic electron-optical effects are most clearly recorded
in diamond knife cutting-edges, they have also been observed under
specially controlled conditions in other types of ultra sharp
cutting edges in glass and steel razor edges, as well as other
kinds of related cutting tools. This "ultrasharp knife cutting-edge
electron transmission effect" can serve as a basis for quantitative
studies and selective detection of all types of cutting edges of
these dimensions. After suitable calibration of the transmission
electron microscope image or electron diffraction pattern using
single-crystal graphite or epitaxially grown thin films as a
standard for checking the thickness and related structural
parameters, a conventional electron microscope can be readily
adapted to serve as a routine monitoring and detecting device for
these cutting edges of molecular thickness.
Other similar types of "molecular cutting-edge detectors" based on
the selective electron-transmission affect are suitable for
automated routine inspection and control of ultra sharp cutting
edges on an assembly line basis. Some of these devices, which will
be the subject of a separate application, are considerably
simplified by making use of a collimated electron source (e.g.,
radioisotope .beta.-emitter sources) suitably arranged so that the
electrons selectively transmitted through the ultrathin
cutting-edge are then recorded on a sensitive indicator, such as an
image intensifier or photographic emulsion.
b. Examination of ultra sharp cutting-edges by high resolution
replica techniques.
Ordinary replica techniques which provide an indirect view of a
surface mold or impression on thin plastic and metal films are very
difficult to apply to the study of ultra sharp cutting edges,
because the active cutting edge is largely obliterated or can
actually be damaged during the required stripping process, and
there is considerable distortion of the initial structural
relationships. It was therefore necessary to develop a simple and
reliable replica technique which would yield an accurate impression
of the active cutting edge and adjoining knife facet suitable for
examination by high resolution electron microscopy. Among the
numerous variants that were developed the following "high
resolution indentation-replica technique" applied preferably on
thin films was found to be the most useful method for accurately
determining the cutting-edge sharpness radius, and the surface
detail of the adjoining knife facets, including the orientation and
size of the abrasive particle tracks in the final polishing process
of the ultra sharp diamond knife edges.
It is based on the precise "indentation track" of the active
cutting edge which is produced by perpendicular indentation of the
knife on ultrathin (about 50 to 200 A.) carbon-plastic films
floating on a liquid surface or supported on a smooth soft support.
The atomically smooth and perfectly plane films are produced by
high-vacuum evaporation of carbon on a freshly cleaved mica
surface, followed by coating with a collodion or Formvar film. The
composite films are then stripped off from the mica by simple
floating on water, or glycerol, or on gelatin or plastic
substrates. The diamond knife or other type of ultra sharp cutting
tool is mounted on a special holder connected with a sensitive
balance to permit controlled ultra-low-load identation of the
smooth surface film, when the edge descends perpendicular on this
ultrathin surface. The resulting indents (of which several can be
made in rapid succession without damaging the sensitive knife edge)
are readily visible with phase-contrast or dark-field microscopy.
However, the actual active cutting edge can only be seen by direct
examination of the replica (with subsequent carbon or platinum
shadow-casting if necessary) by high resolution electron
microscopy. The cutting-edge sharpness radius stands out clearly
and can be accurately measured, allowing for the contribution of
the shadowing material. Adjacent to this cutting edge replica, the
individual crystalline unit layers of the diamond facet can be
clearly discerned. Profiles of the cutting edge can also be studied
by means of stereoscopic pairs recorded with a special tilting
state in the electron microscope. The replica technique can be
further refined by carrying out the whole procedure in an ultrahigh
vacuum chamber (10.sup..sup.-7 to 10.sup..sup.-9 mm Hg) indenting a
freshly prepared ultrathin carbon film evaporated directly on a
cooled, clean glycerol surface, followed immediately by carbon or
carbon-platinum shadow casting of the resulting indents of the
ultra sharp cutting edges.
Large number of measurements performed on these replicas clearly
show that the active cutting-edge sharpness radius of diamond
knives are exceptionally regular with dimesions of the order of 10
to 30 A. in the best knives. Moreover, these active cutting edges
are remarkably stable after prolonged used, as determined by
combined application of the replica and the other measurement
techniques described here.
There variants of the replication techniques include direct
evaporation of NaCl-carbon films on to the suitably oriented
diamond or other ultra sharp cutting edges in an ultrahigh vacuum,
followed by stripping off in an ultrafiltered water surface.
"Shadowgrams" of the ultra sharp cutting-edges can also be obtained
by collimated heavy metal shadow-casting of the cutting tools
resting on a freshly cleaned mica surface.
c. Determination of properties and stability of ultra sharp cutting
edges by cutting of ultrathin serial sections of metals and
selected specimens, followed by examination of the sections and
surface of the specimen block.
All of the previously described methods give valuable data on the
dimensions and configuration of the ultra sharp cutting edges, but
the most critical test for their stability and performance can only
be provided by actual ultrathin sectioning. For this purpose a
special instrument, such as the ultramicrotome described in my U.S.
Pat. Nos. 2,961,908 and 3,091,144, preferably operating at very low
temperatures, is employed. Metal specimens, such as single-crystals
of aluminum, copper, gold, zinc, or platinum with a shaped "pyramid
or cone" tip, preferably only 10 to 100 microns in diameter, are
mounted on the specimen holder of the very accurate ultramicrotome
with a continuously adjustable feed, and a measured
reproducibility. c. Determination of properties and stability of
ultra sharp cutting edges by cutting of ultrathin serial sections
of metals and selected specimens, followd by examination of the
sections and surface of the speciment block.
All of the previously described methods give valuable data on the
dimensions and configuration of the ultrasharp cutting edges, but
the most critical test for their stability and performance can only
be provided by actual ultrathin sectioning. For this purpose a
special instrument, such as the ultramicrotome described in my U.S.
Pat. Nos. 2,961,908, and 3,091,144, preferably operating at very
low temperatures, is employed. Metal specimens, such as
single-crystals of aluminum, copper, gold, zinc, or platinum with a
shaped "pyramid or cone" tip, preferably only 10 to 100 microns in
diameter, are mounted on the specimen holder of the very accurate
ultramicrotome with a continuously adjustable feed, and a measured
reproductibility without the specimen of about .+-. 10 A. After
careful adjustment of the optimum cutting angle of the ultra sharp
(diamond) knife, keeping the clearance angle as small as possible
(preferably less than 1.degree.) the thermal specimen advance is
activated, and preferably uniform serial sections are cut and
collected in the form of a tenuous ribbon floating on the water (or
other suitable liquid surface for low-temperature work) contained
in the trough. Metal sections are ideal test specimens for the
ultra sharp knives, because in contrast to the usual
plastic-embedded specimens which have a tendency to "spread-out"
sectioning, sectionig, the metal sections retain their original
dimensions, and give a faithful reproduction of the diamond cutting
edges, glass and steel knives cannot be used for ultrathin
sectioning of hard materials, such as germanium. It is important to
bear in mind that the process of cutting a thin metal section is
accompanied by a rather severe shear deformation of the section and
a momentary rise in temperature, which can be as high as
300.degree. C when sectioning is performed at room temperature.
However, when ultrathin sectioning of metals is carried out at
temperatures below about -180.degree. C this deleterious
temperature rise can be largely avoided (because of the typical
high termal conductivity, low thermal expansion and low
compressibility of diamond).
The resulting ultrathin sections of metals are about 50 to 200 A.,
as determined by combined electron microscopy and electron
diffraction studies, using control films and crystalline lamellae
of known thickness for accurate calibration. The structure of the
metal sections is a very sensitive indicator of the dimensions and
quality of the ultra sharp cutting edges, since a "blunt" knife
will result in section failure, edge deflection will result in
irregular and compressed sections, etc. The corresponding block
surface from which the sections have been cut exhibits an
exceptional degree of finish (in the microinch range) which is
superior to that obtained by any of the known mechanical polishing
procedures. Nevertheless, even slight imperfections in the cutting
edge show up as minute "tracks" in both sections and block
surface.
One of the most sensitive criteria for determination of edge
thickness and quality is provided by the preparation of ultrathin
serial sections of macromolecular components and highly ordered
biological tissues with a precise "built-in period" which serves as
an internal calibration for the section thickness. Thus, an
ultrathin section of about 90 thickness as revealed in an electron
micrograph of the highly regular paracrystalline lattice of insect
wing inclusions depicting individual protein molecules with a
regular spacing of 80 to 90 A. It is actually possible to section a
virus particle, like the tobacco mosaic virus, of molecular
dimensions into two or more than 80 A.
Finally, by using the best ultra sharp cutting edges and
low-temperature ultramicrotomy, it has been possible to produce
serial sections having molecular or sub-molecular thickness,
prepared from frozen macromolecular substances. A native crystal of
the enzyme catalase with a regular spacing of 90 A. has been cut
into sections which are about 100 A. thick; another catalase
crystal has been sectioned into slices of about 45 A. thickness.
This can only be directly visualized by high resolution electron
microscopy when working with extremely thin sections of these
molecular dimensions.
An even more sensitive criterion for cutting tool sharpness can be
obtained by sectioning long-chain polymers such as starch into
their constituent sugar molecules, as mentioned previously. This
true "molecular sectioning" results in characteristic chemical
changes (such as the different specific chemical reactions of sugar
molecules, prepared by controlled breaking up of the starch
molecules) which can be correlated with the known structural
parameters of these molecules, and in turn related to the
cutting-edge sharpness of the knife.
It is important to emphasize that there is a very close correlation
and excellent agreement between the results of the different
methods described here for determination of the dimensions and
properties of these ultra sharp cutting edges.
Apparatus for effecting ultramicrotomy at low temperatures to
produce serial sections of long-chain polymers and other important
macromolecules into their active constituent components is
illustrated in FIG. 4. Because the diamond knife used in
conjunction with the cryo-ultra-microtome, as shown in FIG. 4, can
cut specimens as thin as 10 A. to 100 A., it is now possible to
literally effect chemical changes by cutting. Thus, by means of
molecular sectioning a starch molecule can be cut up in such a way
that it becomes sugar. More significantly, it is now possible to
cut the master molecule DNA into viable segments which can be
subsequently spliced onto other DNA molecules.
In the apparatus shown in FIG. 4, a diamond knife 1 of an edge
thickness 0.001 to 0.01 micron is attached to a special holder 3,
provided with adjusting screws 2 and can be cooled down to
temperatures of about 1.degree. to 30.degree. above absolute zero
by cryogenic fluid supplied through a tube 4 to a cavity in the
holder to provide a bath for picking up the ultrathin serial
sections. The holder can be precisely adjusted by a micrometer 5.
Lateral movements of the holder can also be controlled by a screw
6. A flexible cooling strip 7, for example copper, ensures the
required low temperature which is critical for biological
specimens. The entire assembly base 8 is filled with a cryogenic
fluid such as liquid nitrogen or helium through a flexible tube 9.
The specimen 10 is affixed to a holder 11 which is also cooled
through a device 12 and is attached to a highly precise cylindrical
metal rotor 14 which is kept cold through special cooling coils 13,
15. A conventional motor drive operating in a high vacuum can also
be attached to the rotor at 13, 15. Special diamond or equivalent
point bearings 20 maintain the constancy of the rotor ends 16.
Provisions are made by means of copper or other cooling strips 17,
18 to keep the bearings at the required low temperature. The
bearing holders 20, 21 and 22 are needed for extra stability. The
entire ultramicrotome is preferably enclosed in a high vacuum
enclosure 24 with seals 23. An ultrahigh vacuum can be maintained
in this enclosure by using an ion pump 25 in conjunction with
cryogenic pumping. Since the system is very sensitive to
temperature changes, radiation shields 27 are provided. The rotor
can also be driven by a special cryogenic magnetic motor drive 29
comprising pick off pins A, an upper bearing B, a stator shell C,
rotor D and lower bearing E. The entire assembly is mounted on a
vibration proof case 26.
With the cooling means illustrated and described, it is possible to
cool the specimen and its holder and the diamond knife and its
holder and also the whole environment of the specimen and knife
down to a temperature of the order of 0.001.degree. above absolute
zero. By cutting with a diamond knife having an edge thickness of
0.001 to 0.01 micron and by carrying out the cutting at a
temperature at or below 20.degree. K (the temperature of liquid
hydrogen) it is possible to obtain much thinner cuts than
heretofore possible. Preferably the cutting is effected at a
temperature below 50.degree. K. While liquid helium provides a
temperature of 4.2.degree. K, it has been found possible to achieve
still lower temperatures by using helium 4 and helium 3 isotopes,
preferably in dilution type refrigeration.
The adjusting screw 2 and micrometer 5 are used to effect accurate
initial positioning of the diamond knife relative to the specimen
carried by the holder 11 on the rotor. Means is incorporated in the
mounting of the specimen holder for moving the specimen toward the
knife holder by minute increments to cut successive ultrathin
slices of the specimen as the rotor rotates. Such means may utilize
the thermal expansion of a metal member, for example a member of
INVAR alloy. The thermally expansive member may be sufficiently
heated by light shining on it. Preferably, however, movement of the
specimen is effected by magnetostrictive means. Thus, the specimen
mounting may comprise a bar or other member of the rare earth
dysprosium which deforms physically in response to a magnetic
effect.
The described cryogenic ultramicrotome can be sealed down (as shown
in the scale) from 50 to 1 centimeter approximately. The molecular
or pauciatomic method has been thorougly tested and can be used to
section any type of material ranging from biological specimens to
metals, catalysts etc. This type of "bio engineering" which has
been made possible by the introduction of the diamond knife, has
numerous other applications extending into the micro electronic
industry (e.g., production of micro coils and micro transformers or
capacitors by precision sliding). The key feature of this approach
is given by the capabilities of the electron microscope which not
only make it possible to see directly the critical domain of atoms
and molecules but also to be able to "design" matter in this
pivotal range which was hitherto inaccessible.
d. Examination of ultra sharp edges by ion-optical, field emission
and electrical measurement techniques.
One of the most interesting analytical approaches in the study of
ultra sharp cutting edges, particularly of diamond, which is being
actively investigated by the applicant, derives from the
application of point-projection microscopes. This type of
microscope, as first described by E. Mueller, consists of a sharp
metal cathode (e.g., very fine tungsten point with a radius of only
a few hundred Angstroms) located in a highly evacuated or
gas-filled tube with the fluorescent screen and an interposed anode
ring at a specified distance. With a sufficiently high potential
(usually of 10 to 20 kV) between anode and point, electrons or
ionized helium atoms will be drawin from the point and imaged
directly on the screen displaying the atomic structure of the point
at magnifications of several million. The magnification can be
readily calculated as a function of the radius of the cathode and
the distance to the screen. A diamond ultrasharp edge of 10 to 100
A. which is coated with tungsten or platinum by high vacuum
evaporation can be actually considered as an extended point source
in such a point projection microscope. Experiments demonstrate in
fact that such a diamond knife edge is an excellent point source
which gives exceptionally bright images both in the field emission
and in the field ion microscope. From these images the dimensions
of the emitting area which correspond to the active cutting edge of
the knife can be determined. Moreover, as shown by the classic work
of E. Mueller, this technique permits in principle a very accurate
determination of the atomic structure of these emitting tips,
investigation of crystalline defects and other important
properties.
Systematic work in this field combined with investigation of the
characteristic elecron-optical and electrical field properties
exhibited by ultra sharp stable edges of these dimensions are of
great potential value for atomated inspection of ultra sharp
cutting edges of all kinds.
A new type of source for electrons, ions, X-rays, neutrons of
controlled exceptionally high intensity and coherence is
illustrated in FIG. 1.
A diamond point or edge 30 having a tip radius of approximately
0.001 to 1 micron or up to about 10 microns and of a precisely
determined shape acts a permanent substrate for sources of a
suitable material such as tungsten, rhenium, lanthanum, barium,
caesium and other related materials of suitable work function and
physical properties for electron emission. As shown in drawing FIG.
1, the diamong tip or edge is accurately mounted on a suitable
ceramic or tungsten base 31 which is mounted on a ceramic base
plate 32 so it can be precisely aligned. For example the base 31 is
provided with a reduced threaded portion 31a which extends through
an upper washer 33, a hole in the base plate 32 and a lower washer
34 and is secured by a nut 35. The hole in the base plate is
sufficiently large to permit adjustment of the base 31 carrying the
diamond tip whereupon the nut 35 is tightened to hold the diamond
tip in adjusted position.
Associated with the diamond tip 30 is a heater and emitter 36 in
the form of a strip or coil or other suitable configuration. The
emitter-heater is shown by way of example as a strip extending
between two posts 37 and having a hole through which the diamond
tip extends. The strip 36 and also the posts 37 are preferably of
tungsten or rhenium. The tungsten or rhenium strip 36 is heated to
a high temperature by current supplied through the posts 37 in
order to evaporate a thin film of this emitting material onto the
diamond point.
The assembly is placed in a "cathode gun" (see FIG. 1) of suitable
shape and material and serves as an electron source which, in
conjunction with an anode (not shown in drawing) is used to
accelerate an electron beam of very high brightness, for example,
10.sup.4 to 10.sup.6 amperes cm.sup..sup.-2 Sterad.sup..sup.-1 as
shown in FIG. 2. As illustrfated in FIG. 1, the gun includes a
cathode cap 38 of stainless steel or molybdenum having a molybdenum
member 29 with an aperture through which the tip of the diamond
projects. The aperture is of a size to receive the diamond tip
closely without touching it and may, for example, be of the order
of 0.1 to 1 mm. The cap shields the anode from emission by the
strip 36 and thereby assures that only the diamond tip emits. A
bias voltage of, for example, 100-1,000 volts is applied between
the emitter 36 and the cap 39 while an accelerating voltage of, for
example, 1,000 to 100,000 volts is applied between the cap and
anode.
In order to increase the life of the cathode, a reservoir of the
emitter is provided. This may, for example be in the form of
emitter material preferably in paste forms introduced into a
transverse hole 30a in the diamond tip or a ring of said material
around the base of the tip as illustrated at 30b. The reserve
material gradually migrates to the hotter tip as the emitter
material on the tip is used up.
The characteristic beam current obtained with a cathode according
to the invention in conventional high vacuum tubes, for example
10.sup.-.sup.4 to 10.sup.-.sup.6 mm Hg, used in electron
microscopes and in television tubes and related devices is orders
of magnitude higher than the normal thermionic sources. The unique
advantages of this type of source are (a) permanent critical
alignment since the diamond which acts as a substrate is not
affected and the source can be readily replenished simply by
evaporating a new film onto the diamond tip. Since diamond has the
lowest thermal expansion coefficient and is the hardes substance
known, it will withstand the great stresses normally involved. The
expected life of cathodes in accordance with the invention is
months or years, as compared with the average lifetime of the usual
cathodes measured in several hundred hours only. This source can be
applied to replace the existing cathode sources for electron
microscopes, television tubes, X-ray tubes, etc. without
appreciable modifications. In view of its exceptional stability and
precise alignment, for example in an electron microscope where
alignment has to be adjusted every time the tungsten filament
source is replaced, it is particularly suitable for applications
such as space probes, etc. requiring months and years of unattended
use. The diamond tip may be a point or an edge according to the
application of the cathode.
The thin film of emitter material on a diamond tip substrate in
accordance with the invention permits critical centering and height
adjustment of the emitter relative to the cap to assure stable bias
voltage and emission control. This electrode and cap design is
applicable to both cathode and anode structures, combining optimum
micro-geometry and operational conditions to obtain
space-charge-free emission and significant field enhancement. The
emitter in accordance with the invention may operate as a
thermionic emitter or as a temperature-field emitter.
A similar type of source as shown in FIG. 3 is used in a field
emission electron and ion tube using either a diamond point or a
diamond knife edge 40 with a tip radius of about 0.01 to 0.001
micron serving as the substrate for accurately deposited tungsten
or rhenium thin film strips, and working in an ultra high vacuum,
for example of the order of 10.sup.-.sup.8 to 10.sup.-.sup.12 mm
Hg. This type of source becomes the equivalent of a "comb type
source" representing many hundreds or thousands of individual field
emission point sources.
Alternatively, the diamond tip may have a continuous coating. The
tip 40 is mounted on a holder 41 shown in the form of a rod
extending down through a vessel 42 cooled by a cryogenic fluid, for
example liquid hydrogen or helium. The diamond tip 40 is surrounded
by an emitter Ch of a suitable emitter material, for example
tungsten or rhenium. The point of the diamond tip extends through a
hole in a grid cap K, for example of molybdenum. A second grip cap
W having a larger aperture is interposed between the diamond tip
and an anode A.
The cathode-anode assembly is surrounded by an annular chamber 43
which is likewise cooled by a cryogenic fluid, for example liquid
nitrogen, and is enclosed in an envelope in the form of a tube 44
in which a high vacuum is maintained. Shielding is provided by
conductive coatings 45 and 46 connected to ground. A flourescent
tube 47 is provided at the end of this tube. It has been
demonstrated that with an emitter of this kind one can obtain
enormous current densities, for example, of the order of 10.sup.8
amp./cm.sup.2, thus exceeding other conventional cathodes by a
factor of a million. For example, by using this type of source in
the T-F mode (temperature and electrical field emission) with
accelerating voltages of about 10,000 to 200,000 or more, these
point shaped cathodes operating in parallel can yield peak powers
of 3MW or more corresponding to energies of 10.sup.8 to 10.sup.15
ergs or more in microsecond bursts. Further modifications of this
type of source for producint high energy and highly coherent
electron beams, X-rays, neutrons, etc. may be made.
It will be understood that features of the embodiments illustrated
in the drawings and herein described are mutually interchangeable
in so far as they are compatible and that many modifications in
size, materials and specific configuration and design may be
made.
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