XN/Pol vs BF

Posted on June 1, 2015 by vade mecum microscope

Pardon?

If the cryptic title of today’s post seems bizarre please remember that for most people microscopy is limited to only one sort: light microscopy. Further one could say that for most people light microscopy is limited to one particular sort: bright field. For the last few posts we’ve been getting into polarized light microscopy in a very general way and with that in mind the obscure little title should make perfect sense.

XN is still used in some papers by authors who wish to refer immediately to crossed Nicol prisms, or crossed polaraizer and analyzer. In practice one may rapidly find the point at which their polarizer and analyzer are crossed by turning one or the other until the light which is seen to pass between the two is at its lowest ebb.

XN in Action

The following photomicrographs were all taken with trinocular AO Spencer microscope using a Nikon 1 J1 consumer grade digital camera fitted with a Nikon 1 to c-mount adapter and c-mount to 23.3mm microscope eyepiece tube adapter. The polarizing apparatus was constructed out of two small discs of polarizing film which cost only $5.00 with shipping. If a full sized image is desired one need only click the image but be warned they are several MB in size. First will be shown the specimen in bright field, followed by the object under XN.

A human hair.

The same hair.

A mouse hair.

The same hair.

Portion of a fly wing.

The same flys wing.

A centipedes forcipule.

The forcipule again.

Many natural fibers, everything from cotton to the hair on ones head, are strongly birefringent. Different forms of fiber will show differently under crossed pols. A motivated individual can discern much from a hair without resorting to such destructive methods as scale casting.

Insects may be surprisingly dull subjects for polarized light and the process may do little but reveal how much dust was remaining on the specimen, as in the case of a hastily mounted flies wing which was collected from a disused attic. At times one may find with surprise that small portions are powerfully birefringent, as in the case of the hardened, venomous, forcipule of the garden centipede pictured above. Frequently only the mouth parts of a specimen will show double refraction. This simple fact can be immensely helpful when trying to identify the mouth parts accurately in whole mounts, especially when optical sectioning is insufficient.

One of these days I’ll have to put up a video of some chemical crystals under XN, stay tuned! -K

Elucidating Illuminators: I

Posted on April 7, 2015 by vade mecum microscope

Integrated Illumination

The choices one has in microscope illumination are often, of late, made by the microscope manufacturer. That choice is not always the one that the user would make for themselves, or even the best for general use. In general one might say that with integrated illumination, whatever the quality, age, or price of the microscope, one can rely on prompt success when it comes to simply obtaining an image. The perfectness of that image may range from excellent to abyssal, but something will certainly be seen at the eyepiece.

Today, if one is possessed of a microscope with integrated illumination little need be done for acceptable operation. A microscope equipped with a mirror is as easy for general, but more complex for critical work, when compared to a stand with integrated illumination. The mirror microscope is next to impossible to use at its full ability without a measure of effort and understanding of some of the methods of illumination for transmitted light microscopy. Let us begin by looking first towards a number of microscopes equipped with integrated illumination of various sorts and speak to the merits and abilities of each.

Köhler Illumination with the BalPlan

This model of the Bausch & Lomb Balplan uses a halogen lamp mounted in the base to illuminate a ground glass Plano-convex lens secured in the base. From there, a beam is sent through an iris diaphragm and fixed focus lens towards a right angle mirror which sets the light vertically through a second lens. The coherent light which emanates from this lens is then passed through the microscopes rack and pinion 1.25N.A. aplanatic condenser. The condenser is equipped with an auxiliary lens which may be swung into place to expand the pencil of light and provide a large enough light source for use with a low power finder objective.

With an object in place (and in focus) it is the work of a moment to obtain a focused image of the iris of the illuminator using the condenser, and adjust the iris of the condenser to so that the numerical aperture of the condenser matches that of the objective for true Köhler illumination. The method of August Köhler to provide a field of light perfectly uniform and out of focus, while retaining all the characteristics required for maximum resolution are today recognized and accepted as optimum for advanced work. Köhler illumination puts a focused image of the light source (the filliament of the lamp or grain of a ground glass screen) at the iris diaphragm opening of the substage condenser and again at the back focal plane of the objective. A bright evenly lit field is then formed at the iris diaphragm opening of the illuminator and in the image plane of the focused microscope slide.

In the photograph the illuminators iris diaphragm has been stopped down well beyond what would be used in practice to better show the in focus diaphragm simultaneous with the image plane of the specimen. Note that despite the light source being a ground glass there is no visible grain in the photomicrograph.

Critical Illumination with the AO Fifty

This American Optical Fifty series microscope makes use of an uncomplicated 15watt (medium base) incandescent bulb as the light source. Mounted in an adjustable housing in the microscopes base the light of the bulb is sent first through a blue filter. Immediately above the filter is mounted a single short focus Plano-convex lens with a ground glass flat surface. This arrangement makes up the integrated illuminator common to microscopes of the Fifty, Sixty, One-Fifty, and One-Sixty series. Above the field lens of the illuminator is a 1.25N.A. Abbe condenser on rack and pinion adjustment.

With this setup Köhler illumination is not possible. Instead a less perfect but far less mechanically demanding type of illumination known as critical illumination is possible. Prior to the modern^∗ acceptance of Köhler illumination as the gold standard it was practice to work with a large light source and less complex illumination apparatus. Rather than a focused image of the light source, a brightly lit field is formed at the opening of the substage iris diaphragm and the back focal plane of the objective^†. Unfortunately, this results in a focused image of the light source (and all its attendant grain or irregularities) being formed in the image plane of the specimen. The worker is then apt to throw the substage condenser slightly out of focus so as to avoid the distracting image of the filament when not using a sufficiently large homogeneous light source.

Not that in the upper photomicrograph there is evidence of dust on the ground glass illuminator, as shown by dark areas indicated by the pointer. In the lower photomicrograph the substage condenser has been thrown slightly out of focus to prevent grain and dust from appearing in the image plane of the specimen. In general use one may chose to rack the condenser up or down if the grain of the illuminator is found distracting, there is however a slight increase in spherical aberration and decrease in resolution.

Add-on Integrated Illumination

Largely as a response to the need for simplicity and convenience when introducing microscopy to students, a large number of manufacturers began to sell small self-contained light sources in the mid-twentieth century. These simple lamps could be used in the conventional way with a mirror bearing microscope by sitting the lamp upon the table, or by mounting it directly to the base of the microscope. The Bausch & Lomb lamp shown here was available with two different mounting brackets; one which made use of the mirrors mount, and that pictured which would be fit over the removable substage iris diaphragm before it was screwed back into the base of the substage condenser.

This illuminator consists of a small 15watt (medium base) incandescent bulb in a switched, sturdy, Bakelite shell. Over the bulb is mounted a blue glass filter with one ground glass surface, followed by a Plano-convex lens. The exposed convex surface of the lens is ground as well. Depending on the arrangement in which the illuminator is used, one can work with critical illumination or not, but again Köhler illumination is not possible^‡.

Such lamps are chiefly recommended by their simplicity, durability, and availability at low cost. Less often mentioned is the versatility of such lamps. Any illuminator of this sort^§ may be arranged to provide ample light in most circumstances a beginning microscopist is likely to encounter. It may be used with or without a substage condenser, or even without the mirror; as it may be set on the table between the foot of the microscope with the light projecting upwards through the stage. If such a light source is to be used it should be considered a component of the microscope, and kept with it at all times.

In the upper photomicrograph the lamp was placed on its back below the substage. Although the illumination is not critical the central portion of the field is even lit and free of grain. In the lower photomicrograph the lamp was positioned ten inches from vertical axis of the microscope and its light reflected by the plane mirror. With the surface of the ground glass focused critical illumination resolves the center of the field of view with slightly less spherical abbaration, although grain is once more introduced into the image and it is evident that this author did not take time to properly center the lamp.

Notes:

∗Well into the twentieth century arguments continued to be raised as to the technical or practical superiority of the various methods of specimen illumination. One may at first wonder at the controversy but it seems to have largely resulted from the need for more specialized light sources with Köhler illumination and the practicality of critical illumination when used with any large (flame or cloud for example) light source. The past popularity of point light sources is a product of the growing pains of the adoption of Köhler illumination as it was generally felt that a bright point of light provided more capacity for filtering without diminishing intensity.

†Technically, in critical illumination that brightly lit filed is formed just before the back focal plane of the objective and moves closer towards it as the power of the objective is increased.

‡Obviously with the lamp mounted directly to the substage condenser one can not bring the ground glass surface into focus in the specimen image plane as is required for critical illumination. Working with the lamp on the work table, or with the lamp mounted to the mirror bracket it may be brought into focus with the substage condenser if it is in range of the primary focus. I am uncertain as to why B&L would grind the convex surface of the lens, but I suspect it was motivated more by the desire to make the device less open to damage by unskilled hands (there are no exposed polished lenses to worry about) than by optical concerns.

§All of the major manufacturing houses put out similar lamps with differing arrangements as to bulb and glass. A popular American Optical model was equipped with a blue filter and swing out ground glass or bullseye lens, providing a limited means of controlling the intensity of the light. Modern LED or incandescent lamp housings are available that will mount in place of the double sided mirror of most vintage stands, one need only verify that the mounting is compatible.

If things seem complicated don’t wait on the availability of fancy equipment, great work has been done with desk lamps and white clouds. Next time we’ll look to external illuminators. -K

Digital Photomicrography with the Student Microscope

Posted on March 10, 2015 by vade mecum microscope

I can not stress enough the importance of traditional micrography as a means of gaining understanding of the specimen, but for ooh and ahh factor digital is king. -K

The setup

For digital photomicrography a consumer grade mirror-less digital camera with removable lens is mounted over the eyepiece with a two part connector. The first part fits into the cameras lens socket and is friction fit to the second part which fits over the microscopes tube and rests upon its shoulder. The camera may be removed at any time without difficulty and all apparatus is away from the ocular so that it is un-obstructed and may be changed. With the camera removed the microscope may be focused as normal without recourse to the cameras display. As the imaging sensor is at the microscopes eyepoint no secondary focusing is required when the camera is moved into place.

The microscope is that same common workhorse that has been featured in the previous entries. Lighting is provided by a single 60 watt cool-white incandescent bulb in a goose-neck desk lamp. A 5x Huygenian ocular is used for every image. The camera is an older Nikon 1 J1. Anyone desiring to know the settings used for each exposure is advised to check EXIF data for each image. A traditional test object, the proboscis of a blow fly, is used for each image. Be warned, clicking on the images will display a full size (3872×2592) image of several megabytes size.

Photomicrographs

First we remove the lower portion of the divisible 10x objective leaving a perfectly serviceable 32mm equivalent focus objective. A larger aperture opening is spun into place with the circular diaphragm to avoid vignetting the image.

Not bad considering the aberration inherent in such a lens. Observe that despite being relatively close to the center of the field of view the finer points of tung are out of focus.

***

With the assembled 10x divisible objective (16mm EF 0.25NA) spherical aberration in particular is much less obvious. A smaller opening of the circular diaphragm is selected to provide better contrast and reduce glare. No realignment of the concave mirror was made.

Much more of the structure of the tung may be made out although the depth of field is noticeably lacking. Only the smallest evidence of chromatic aberration is visible. Despite what is a very rudimentary lighting system the field is bright and even.

***

Switching to the 43x (4mm EF 0.65NA) objective we switch also to the smallest available aperture in our circular diaphragm. Unlike the previous change in magnification using the divisible objective the change from 10x to 43x is relatively parfocal and a few turn of the fine focus results in the following image.

The field is surprisingly well lit for the absence of a condenser. One may observe that depth of field is more than one might expect especially when considering the relative thickness of the specimen. The color fringes of chromatic aberration are in evidence and anyone accustomed or intending to do much work at this magnification would surely be unsatisfied with the image. Understandably, one with experience might forget that those relatively new to the pursuit are less sensitive to such things and would likely be rather happy with the quality of the above image.

***

Turning the arm of the mirror sharply to one side and switching to the largest opening of the circular diaphragm we are able to take advantage of an all but forgotten lighting technique. A shade is placed so that the area below the diaphragm opening rests in shadow and the lower portion of the divisible 10x objective is removed.

Oblique lighting is not generally possible with a substage condenser. Specialized stops may be put into place but even then the obliqueness of the light is subject certain limitations relating to the working distance and numerical aperture of the condenser. Here a starkly black background is visible because the surface of the table is black and out of focus for the objective. The specimen is brightly lit by the mirror and that light which it sends into the objective. Every speck of dust on the slide is noticeable and in the full size image one has no trouble at all in identifying to which side the mirror was swung (to the left).

***

With the fully assembled 10x objective a passable image is produced which is serviceable as a “poor mans dark-field”. Patch stops intended to render a standard condenser a dark-field substitute do not provide so dark a background.

The Camera Lucida

Posted on April 11, 2014 by vade mecum microscope

Lets toss this off quick and move on to something else… -K

The camera lucida is a sort of artistic crutch which relies on the dedication of its adherents for its success. To be sure one can enjoy incredible results with the camera lucida, but the requirements of its use coupled with the complexity and expense of the device are sufficient that for casual micrography one is better off with other methods. For the individual with the rigid dedication to acquire skill with the camera lucida it is a tool of wonderful capability, for nearly everyone else it’s a device of torture.

The principle on which the camera lucida operates is reflection. Whereas in the previous post an image of the specimen was reflected onto the drawing surface, here an image of the drawing surface is reflected into the eye as it simultaneously observes the specimen. Apparatus which achieves this has been adapted to all manner of situations and some schools of art teach the use of the camera lucida to artists as a method of perfecting perspective. For micrography the camera lucida seems to have fallen largely by the wayside, perhaps even more so than micrography in general. However, modern camera lucidas are still available and there are occasional adherients in professional and amateur microscopy who make regular use of vintage instruments.

Idealy one wishing to begin serious micrography with the camera lucida should purchase the apparatus. If however one wishes to get an idea of the use before making the expenditure a simple experiment can be made with nothing more than a coverglass. Incline the microscope so that the body is horizontal and position a coverglass at a forty-five degree angle over the eyepoint of the ocular. When aligned correctly and the specimen and area below the coverglass are illuminated just right one will see both when the eye is placed close above the coverglass.

Apparatus of this sort was manufactured for many years which consisted of little more than a coverglass (or neutral tint reflector) in a frame held to a circular fitting which would slip over the rim of the ocular. Professor Abbe improved on this design significantly (others did as well but Abbe’s apparatus is the most generally found) by replacing the simple reflector with a neutral tint reflector having a small hole in its center and cemented between two right angle prisms. Parallel with the plane of the reflector he placed a mirror which would reflect the image of the drawing surface onto the reflector. With the device over the eyepoint of the microscope ocular one could observe drawing surface and specimen as a single image while keeping the microscope inclined in the usual position.

Excepting skill, success with the camera lucida is largely a matter of alignment and lighting. Proper alignment is had by first ensuring that the position of the reflector is in the plane of the oculars eyepoint. If this condition is not met no description of the difficulty and aggravation written here will serve. Next one must position the microscope so that the drawing surface is in plane with the image provided by the eyepiece. In practice this is most easily achieved by keeping the microscope fully vertical and the drawing surface fully horizontal. If this is the case the mirror of the camera lucida may be positioned at a forty-degree angle. If one has a camera lucida with the mirror fixed at an angle other than forty-five degrees the drawing surface will have to be inclined to ensure that a ray of axial light is at a 90 degree angle with the drawing surface.

Once things are aligned lighting must be considered. Light the specimen well enough for clear vision, and the drawing surface well enough that it is not overpowered by the light of the specimen or overpowering of it in turn. The variation of the Abbe camera lucida by Bausch & Lomb pictured below bears a variable series of filters which may be turned into the path of light reflected into the eyepiece to afford some level of moderation. In practice it is often more expedient to simply reserve a variable intensity lamp for illuminating the drawing surface.

The upper portion with filters to limit light sent to the eyepiece reflector.

Below one can see that with the microscope fully vertical the mirror may be positioned at forty-five degrees and the drawing surface left horizontal without distortion of the drawing surface. This is not the most comfortable position for use but it is the simplest to set up. Two illuminators are visible in the photograph. That on the left is used exclusively to light the specimen and bears a frosted daylight glass and 0.3 neutral filter. The illuminator on the right is used only for lighting the drawing surface and bears a frosted daylight glass as well. The bulb in each case is a 75watt Mazda halogen spot lamp.

Abbe camera lucida by Bausch & Lomb circa 1920 arranged for use

Looking into the camera lucida one is presented with an image of the specimen and anything that appears below the mirror. If the image of the drawing surface is too dimly lit it will be impossible to see the image of the pencil point and therefor impossible to trace the outlines of the specimen. One may find that with low powers it is often necessary to dim the light on the specimen and increase the light on the drawing surface. In the case of higher powers one will find the lighting needs reversed. With some varieties of camera lucida one must be certain to keep ones eye in the same position continuously until the micrograph is complete, otherwise the image seems to move about on the paper. With the Abbe camera lucida one may move the eye as the image can only be clearly observed when it is above the hole in the reflector.

Below is something of the image one sees while looking into the camera lucida. At the right one can see a portion of the pencil while to the left one can see the reflection of that pencil simultaneous to the image of the letter “e” slide. The lighter central portion of the image in the eyepiece is not visible in use but is caused by the borders of the hole in the reflector.

Looking into the eyepiece

Some time I shall have to make more of my efforts with the camera lucida fit to read of. Maybe put up some of the little things it permits one to do into words, but for now the camera lucida is rather more trouble than it is worth. Without an inclined drawing table it is uncomfortable in use and provides generally inferior results as it does not encourage one to spend any time improving ones skill.

Projection Micrography

Posted on April 8, 2014 by vade mecum microscope

Taking pictures of my work area makes me feel as if I should tidy up more often. -K

Projection microscopy can be a means to many ends. One might use it for group demonstrations, measurement, specimen comparison, even casual viewing if one is so inclined. Of course it’s looked at here for the purposes of micrography, to which it is particularly well suited. Micrography which is aided by projection of the image onto the drawing surface is considerably easier than many of the other methods to acquire. It is also among the more inexpensive methods though very much reliant upon the conditions of ones work area.

In the previous methods looked at one required only the usual set up and a skill at drawing with perhaps a graticule for assistance. In projection micrography one may be excused for initially thinking that a projection microscope is required, but that is just not the case; one can get by with little more than a microscope. Naturally there are bits of equipment than can simplify things, specialized accesories and specialized microscopes one can purchase, but if one is without funds to do so, or just eager to try projection micrography today with what is on hand there is no need to wait.

One will need a microscope which may be inclined so that the eyepiece is horizontal, a powerful illuminator, and paper and pencil. When gazing into the ocular one is presented with a magnified virtual image that is optimally viewed at the eyepoint of the ocular. However, when the eye is beyond the eyepoint of the ocular, or the ocular is removed, the virtual image can still be observed. An other image is produced however, a real image. This image can be thrown upon a screen or sheet of paper simply by placing it in the path of the rays which pass through the objective or objective and ocular. The size (not the magnification) of the image on the screen moderated by nothing more than the distance of the microscope from the screen.

With a traditional microscope one may incline the stand horizontally and using an external illuminator send light directly through the slide without making use of the mirror. If a wall (with a paper affixed to it) is sufficiently nearby, the room is sufficiently dark, and the illuminator sufficiently bright, one may simply focus the specimen by observing the image thrown upon the wall and trace it onto the paper. In some cases it will be helpful to use only objectives and oculars of the lowest power, or to employ only optics which have large object lenses that permit more light to pass though.

Drawing on a vertical surface is rather awkward and one would naturally prefer to have the image thrown onto a table. This is where various specialized bits come into play. Prisms and mirrors can be positioned so as to send the image from the microscope onto a table or wall regardless of the position of the body tube. The simplest sort of device is a mirror that may be positioned at forty-five degrees from the horizontal body tube, and constructed quite cheaply from a ladies compact. To size the image conveniently it may be necessary to place a book beneath the foot of the microscope. Below is an example of the set up which makes use of a mirror made for the purpose by the Bausch & Lomb company. The objective is a 32mm (160mm Tube Length) achromat.

Lange’s is an indispensable resource in any lab!

One might have seen projection oculars available and be tempted to believe that any poor results experienced are the fault of the optics. Before spending the money on specialty oculars (which have their uses) one might observe the images below. The first image is that from a regular 12.5x Huygenian by B&L, while the second is a 12.5x projection ocular by B&L. All else being the same, (except for the steadiness of my hand at the camera) one notices immediately that the first ocular produces a sharper, more tightly constrained image. Why then are projection oculars generally more expensive? Without getting into it, lets just say that they have their uses and under particular conditions to which they are suited they more than justify their expense. The demonstration here is to illustrate that one needn’t have a special set of optics to project an image for micrography.

A 12.5x Huygenian ocular

A 12.5x projection ocular

The image below was made without any ocular at all. One can see that the letter “e” is oriented differently than in the images made with an ocular. If one cares to recall that there are no extra optics in the body of this particular microscope,only the objective and mirror being used to create the image, one can better understand something of optical principles. One should also note the uneven illumination of the field because this was example was not set up for either critical or Köhler illumination. Historically oculars were not used for much projection or photographic uses, the reason was largely related to the apparatus employed but it is worth mentioning that in most circumstances if an ocular is not used one can obtain a brighter image; by using a 40x objective alone rather than with a 4x objective and 10x ocular together for example.

To make it larger I need only have lowered the drawing surface

Why not try projection micrography today! It can be a wonderful way to better understand the optical workings of ones microscope and produce micrographs at the same time. Below is a sketch of the letter “e” slide produced by this technique. As with the previous micrographs the sketch was made in under five minutes. Because of the method used more detail could be put into the image, which is also more precise than any of the micrographs produced previously.

More details and more accuracy, more involved set up too

If one wishes to ensure that the image is accurately projected be certain to use a reflector that is large enough, and near enough to the eyepiece (if one is used), to project the entire field of view. It is then a simple mater to measure perpendicular axes of the projected image to ensure it is circular. As a final word on this method: the darker the work room the easier things will be.

Assisted Freehand

Posted on March 30, 2014 by vade mecum microscope

In a sense it’s all just assisted freehand, but I think the meaning is clear. -K

Yesterday efforts were made at getting something on paper and learning a bit on setting up properly. Regrettably those efforts will have accomplished very little of a immediately practical sort. The images produced might be used to visually identify a slide at a glance if its label has fallen off, but that is assuming there are no similar slides on hand. My little sketch could verify that it is of a letter “e” but one would be hard pressed in putting it definitely to a particular unlabeled slide of a letter “e”. Because there was no assistive device that would help with sizing and dimension, we end up only with a general rather than an accurate depiction; one couldn’t use the sketch to measure the size of the letter.

Fortunately, the technique used previously can be rendered very practical by the addition of a simple object that may be purchased for from ten to fifty dollars, or made at home for a good deal less. This device can be used for any number of very usefull tasks from micrography to counting, measuring, or locating. One can use it on essentially any microscope at any magnification and it will last a lifetime. This miraculous device? The humble graticule also erroneously designated the reticle or reticule.

A graticule, the name of which comes from the Latin for gridiron, is nothing more than an optical device on which a ruling of small squares has been marked. This is technically the only accurate description for a graticule as markings composed of either various lines or any grid which is not broken up into even shapes would be a reticule; a micrometer or sighting ocular is a reticule, while a simple counting ocular is a graticule (excluding of course the complex rulings of Neubauer which would be in places reticule or graticule). In practice few people make any distinction between the terms and one is likely to find suitable graticules under various names.

The use of a graticule is quite simple and requires only an eyepiece that contains a diaphragm located at the point in which a real image is formed. In practice this means a negative ocular of the Huygenian type with a field stop between the object lens and the eye lens. At the optical plane of the field stop a real magnified image of the object is formed, which may be observed by placing a circle of tissue paper or frosted glass thereupon. When the graticule (or anything for that matter) is positioned at a point where a real image is formed it will be seen conjugate with the virtual image one observes when looking into the eyelens of the ocular. For more detail concerning real and virtual images one can consult the section on Object-Image Math in Douglas B. Murphy’s excellent book Fundamentals of Light Microscopy and Electronic Imaging.

So, assuming one posses a graticule either purchased or home made and a Huygenian ocular with removable optics, one may fit the graticule to the ocular. Begin in a scrupulously clean and dust free area, as nothing is quite so infuriating as introducing dust to the inside of an ocular, and unscrew the eyelens. With the eyelens removed insert the graticule which should fit just within the tube without force or slop. For standard oculars by Bausch & Lomb and most American makers a graticule with a 21mm diameter is best. In some cases manufacturers provide split rings (as pictured) which will hold the graticule in place so that it will not rattle about even if the ocular be roughly treated. If one is had and the graticule is to be left always in a particular ocular place the split ring before screwing back into position the eyelens of the ocular.

Ocular body, graticule, friction ring, and eyelens, seen in the order to be assembled.

With the majority of graticules the lines are ruled upon one surface or the other and each orientation should be tried. If the rulings are more clearly in focus with the eyelens screwed fully home then that orientation should be selected before one settles for having a loosely fitted eyelens. With the graticule in place in the ocular one should focus a slide upon the stage of the microscope and observing it with the ocular find both the rulings and the specimen sharply defined. If the rulings will not come to focus conjugate with the specimen, the eyelens must be screwed down or up until they are. The eyelens and orientation of the graticule should be manipulated until the rulings are truly conjugate with the image of the specimen. Which is to say they should be neither more or less sharply defined and appear as if on the same level as the object although overlayed.

For drawing with a graticule one would do best to use an ocular of low power and obtain higher magnification with the objective only. The reason for this is well explained by the equations in the above mentioned book but, suffice to say that the area covered by each square of the graticule at a given magnification will be less when that magnification is obtained with a low powered ocular. With the graticule in place one need only use a drawing surface ruled into squares to transfer by hand the view seen through the ocular to paper in the method described yesterday. The rulings will assist in both positioning and consistently representing the size of specimens sketched.

In the above photo one might notice that the graticule pictured has rulings in only the very central portion. At first it might seem as if the ruled area being less than that of the oculars diaphragm would be a great hindrance as the entire field of view would not appear ruled. The reason for using this particular graticule will be obvious when it is noted that it is not used with flat-field objectives. By limiting the area covered by the rulings to the central third of the field of view one insures that spherical aberration of any significance is not depicted in ones micrographs. Images in which optical defects are faithfully represented are of as dubious value as live blood examinations where quacks point to specks of dust or dirt as proof of one malady or another…

Now then, if one knows the spacing of the rule on both the graticule and drawing surface, together with the magnification employed, measurements of a sort may be taken on the drawing using no more specialized apparatus than a ruler. This method of measurement is rather less precise than others (tough quite accurate) but requires neither a micrometer object slide or micrometer eyepiece and serves well for work of all but the most critical sort.

With a graticule in place and ruled paper one may produce micrographs of impeccable quality and enormous size (by moving the slide so that the contents of a square are transported across the field). If one is of limited resources or has in the past attempted other apparatus without good results, no better aid to drawing with the microscope can be had than the graticule. Unlike other apparatus to be covered later no special positioning of the microscope is needed and no great effort is required to become adept. An ocular fitted with a graticule may be dropped into position and used at a moments notice, I commend it to any microscopist.

Freehand Micrography

Posted on March 29, 2014 by vade mecum microscope

Goodness this is getting to be quite the series, hope it’s not too dull. -K

Whatever method of micrography one settles upon the skills used for freehand will be put to use, by all means take some time with the following even if the intent is to expend funds and effort on more complex apparatus late; don’t put aside freehand as to difficult or simplistic. To better serve as a font of practicality, certain points must be established at the outset. If following along one would do well to use the same microscope, ocular (or series of oculars), and objective (or series of objectives) for every method and apparatus. It’s not so important that they be of the same power as those here employed but it will be a great asset not to later have some question as to what optics precisely were used.

One should use the same slide as well, and for that slide no better may be selected than a permanent or temporary mount of a small letter “e” upon newsprint. It will prove useful as a means of coming to a better understanding of ones microscope, is accessible to all, and comparatively easy object for sketching (although not without the opportunity for additional details). Select the smallest print to be found and mount the letter erect upon the slide.

Drawing freehand from sight is an ability that is quite beyond simple instructions here though all that is needed for it is practice. Instead every effort will be made to set one down the right path to creating micrographs at the outset and skill permitted to develop naturally. However straightforward it may seem, one should not to simply look through the ocular and sketch out an image. Some fine artists may enjoy great sucess in this immediately, but mere mortals would do better to seek out every advantage. First one should consider lighting, not of the specimen but of the work area. Every effort should be made to light the room to an intensity appropriate with that seen through the ocular. For many the optical bench is often well lit which will be found excessively tiresome on the eyes when the long periods of observation required by micrography (particularly at the outset) are spent. It is helpful to use somewhat less illumination on the drawing surface than is had through the ocular.

Ninety years ago one would have made use of sunlight, an oil lantern, or even a 6volt incandescent bulb for micrography. The other options (carbon arcs for example) proving too brilliant or costly even, for high powered work that did not involve a camera. With such sources of light it was often simply a matter of drawing the shades or extinguishing the rooms other light sources; the illuminator providing light for the specimen with enough spilling out to comfortably light the drawing surface. Today those sources may still be used (often in conjunction with various improvised shades), but as ones microscope is apt to include a built in illuminator which is quite effective in limiting light leakage it becomes somewhat more of a challenge to light the drawing surface well. A desk lamp which may be equipped with a dimmer is quite useful if one is without a light source such as depicted. The drawing surface should be comfortably lit so that with one eye looking through the ocular the other may gaze upon it without straining.

An older B&L with a lovely 32mm objective of 215mm tube length.

Which brings out the next point worth making, both eyes should be used. One eye should be always at the ocular while the other, remaining open, should gaze upon the drawing surface. This is essentially the same method one should utilize in operating a monocular microscope, except that instead of being allowed to completely relax the other eye is focused upon the drawing surface. It is something of a tiering arrangement which is why having a well lit drawing surface that is not too bright or dim is so important. If one is accustomed to wearing eyeglasses for myopia they will be need to be worn only if the drawing surface can not be seen otherwise. This is likely to prove inconvenient for those who do not possess oculars of a high eyepoint, though moving the spectacles as close to the eye as possible will often help enough that standard oculars may be used.

For paper one should use a heavy stock of very slight texture. Coarse surfaced paper will prevent one from capturing finer detail when working with sharply defined specimens and light weight papers will not bear sketching well. The plain side of a standard index card is very convenient and easily sorted and stored as well, notes being made on the reverse. Work always with a pencil initially and try not to fear making a mistake. Once the sketch is made it’s a simple matter to go in with ink if it is felt necessary.

Initially one will do well to maintain the drawing surface at the same angle as the stage of the microscope, if not the same level as well. Try this experiment, focus the letter “e” slide with a low power objective and eyepiece and produce two freehand sketches. In the first sketch incline the stage of the microscope as one finds comfortable and use as the drawing surface the table on which the microscope stands. For the second sketch keep the stage of the microscope level and use something suitable to raise the drawing surface to the level of the microscope stage. Produce the sketches rapidly but not without undue care, it should not require more than a minute and only general outlines are needed.

See, no need to go for perfection!

One will find that although their is no special optical apparatus in use (save the microscope of course) the brain processes the image differently when the drawing surface is not at the same inclination as the microscope stage and the sketch in the first instance is rather elongated on the axis of inclination. Additionally, note that although the degree of magnification employed was consistent in each sketch (a 5x ocular and 32mm objective) the first ended up somewhat larger than the second. By making use of each eye simultaneously the manner in which the brain processes the image is such that the first sketch is produced larger to account for the added distance. If one wishes to get into the matter a series of experiments may be made with optics of similar magnification and different equivalent focus (remember most microscope optics produce a magnified virtual image that is seen as though 25cm from the eye) which will prove diverting… but back to micrography.

On the back (generally ruled side) of the cards mark notes regarding the image and the setup by which it was produced. At a minimum include the objective and ocular used and relative position of the microscope and drawing surface. The date and information about the slide would be well included but are not essential, the thing here is to get into the habit of producing micrographs in something of a consistent manner.

One should produce a quantity of quick sketches making use of differing arrangements of microscope and drawing surface until a useful preference is discovered. Some will find that they prefer to have only the table while others may favor some elevated and angled drawing surface. The idea is to get into the habit of using each eye simultaneously and abolish that fear some people have in setting lead to paper. These micrographs are not liable to stand publication but there is no reason to feel anything other than pride in them however they come out, it’s a dying art and any effort in keeping it alive should be commended.

Aside from that consider my quick sketches from above, and the slide from which they were made. On the slide the letter “e” was mounted erect and seen on the stage before me appeared as it would when reading. In the sketch the letter was reversed along both the horizontal and vertical axis. Now in my case I was using a rather simple compound microscope having only the obvious optical components. There are no prisms or lenses hidden away in the body tube and no inclined head to consider. If I were to use an AO Spencer One-Sixty or a B&L Balplan with their accompanying internal optics how might I expect my sketch to differ and what would that mean is going on inside the body of the microscope? It’s all well and good to note the way in which the image moves when the slide is manipulated but a clearer understanding is certain to be had by considering the optics at play in microscopes of differing construction.

Monocular microscopes are of course well suited to this application but there is no reason one should not be able to employ a binocular head if that is all which is available. Simply use only one (that which is on the side of ones dominant hand) of the two ocular tubes and proceed as if a monocular set up were employed. A right handed individual would place their left eye at the right most ocular and view the drawing surface with the right eye.

Next time: Graticules and Huygenian Oculars!