Unit

Microscopically
2 Measurement

2.1 Micrometer
2.2 The Camera Lucida

2.1 Micrometer:
2.1.1 Eyepiece Micrometer:
A microscope can be used not only to see very small things but also to measure them.
Things seen in microscopes are so small that centimeters or even millimeters are too
big. As a result, micrometers (or microns) are used. A micrometer, also written µm, is
one thousand the of a millimeter - its 10-6m. For this, a micrometer eyepiece is used in
place of the standard eyepiece of the microscope. This has a series of numbered lines
inside of it which make it look like a ruler (see image to the right, click on it to see a
bigger version). The images below show what the eyepiece looks like (with its
protective box) and where to put it on the microscope.
Method - How to use it
1) After placing the special eyepiece, it is necessary to calibrate the microscope. To
do this, a calibration slide must be used. This is a glass slide with one one-
hundredth of a millimeter, 0.01mm, engraved on to its top surface (see photo to the
right). Use care when handling this little piece of glass .Since a hundredth of à
millimeter is very small and difficult to see, a circle is drawn around it. This slide
allows us to find out how big things are as we look at them through the microscope
at different powers of magnification. Put the slide on the stage as shown in the
photo. Be sure that the top of the slide (the surface with the microscopic lines
engraved on it) is pointing up.

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2) Set the microscope to low power and focus on the lines engraved on the surface of
the calibration slide. You should see the following:

eyepiece lines

calibration slide lines

3) The number of lines must becounted. As shown in the figure above, the eyepiece
lines have numbers on them where as the calibration slide slines do not. The total
number of eyepiece lines (which will be called X) are from line 21 to 59. That's a
total of 38 lines. The number of calibration slide lines (which will be called Y) show
a total of 10 lines (note that the little lines mark off half spaces and that the first line
is not counted because is shows the zero mark).
4) Calculate how much each line of the eyepiece measures. In other words find out
what distance is shown between each line of the eyepiece. To do so, use this
equation:

5) After calibrating for low power observations, medium power should be calibrated.
Switch to the medium power objective lens and focus once again on the lineset
checked into the top of the calibration slide. Use the same technique as steps 3
and 4 counting the lines and calculating the measurement between two lines on the
eyepiece.

2.1.2 Stage Micrometer:

A Stage Micrometer is simply a microscope slide with a finely divided scale marked on
the surface. The scale is of a known true length and is used for calibration of optical
systems with eyepiece graticule patterns. This is particularly important when alternating
between objectives on one microscope or when using the same graticule in different
microscope.

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 Microscopically Measurement
Typical scale

Unique serial number

engraved here

CS-
Stainless steel
slide mount

100x0.1mm = 10mm

Scale dimensions

Using with a Stage Micrometer:

When used for the measurement of a specimen, graticules normally need to be used in
conjunction with a stage micrometer, this being a standard size 3" x 1" glass slide
engraved with a scale which is placed on the microscope stage. Incidentally, for our
beginner members, note the distinction between micrometer, the scale etched on a
glass disk or on a slide for measuring purposes, and a micrometer. The latter is an
actual unit of measurement. Owners of the Russian MBS-9 and MBS-10 stereo
microscopes are fortunate in that supplied as standard equipment with these
microscopes is a special 8 x eyepiece with dioptric focusing eye lens and with both
scale and grid eyepiece graticules supplied for measurement purposes.
These two particular Russian stereo microscopes, when used with their eyepiece
graticule, do not need a stage micrometer slide since the manual gives conversion
tables to show the value of the specimen which corresponds to one division of the
eyepiece scale or grid at different objective magnifications. In the MBS stereo
microscope eyepieces, the graticule is inserted in the bottom of the eyepiece; this
means that because this position is not a normal field stop, the graticule will not be in
focus when looking through the eyepiece. This is why this is a focusing dioptric
eyepiece; with it you first use the dioptric adjustment to bring the graticule into focus
with the eyepiece, then you use the normal focusing control of the microscope to are
23mm diameter and those with the MBS-10 are 25 mm diameter.

2.2 The Camera Lucida:

The camera lucida or drawing ocular is useful for tracing a magnified image of the
object under microscopical study.
The Camera Lucida does not have anything to do with a camera, but that is what it has
always been known as, and it is an apparatus very popular in Victorian times before the
development of a photographic camera which could be affixed to a microscope. Books

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on microscopy from around 1855 mention the use of photography as an alternative to
the use of the Camera Lucida, but with the later improvements in the photographic
camera with its ease of an instant picture the purpose of the Camera Lucida was
superseded and brought about its demise. However, no matter how good and labour
saving that modern appliances are, there is sometimes some aspect that is not quite as
good as the old thing, and this is to some extent true in this instance. Whilst
photographs are so much easier and quicker in their taking, the Camera Lucida has one
important advantage over the photographic camera, i.e., drawings can be made in
depth.
We all know about the depth of focus, or, as some may prefer to call it, the depth of
field, but whichever way you wish to call it, it is a fact that the higher the numerical
aperture of the objective the more shallow will be the depth of focus. This means that,
when using a high power objective, even small items have to be viewed at different
levels of focus, and this is the drawback of the camera as it can only take a picture of
any one focal plane. The Camera lucida does not have this problem, as drawings can
be done in depth, the different focal planes being brought into view by a slight change
of the microscope focus as the drawing proceeds.
A Typical Camera Lucida:
Camera lucida have a lift-up or slide-sideways top lens unit which at first glance looks
like the eye lens of an eyepiece, but it is more complex than that, for below it is a small
prism with a half silvered surface which (as explained by Conrad Beck in a book entitled
"The Microscope" published in 1938) transmits half the light upwards to the eye from
the microscope and reflects by means of an adjustable mirror on an arm, about 6" to
one side, the light from a sheet of drawing paper placed on the table at one side of the
microscope immediately below the mirror.
The mirror reflects the image of the sheet of drawing paper through a small hole in the
side of the Camera lucida eyepiece cap to the small prism; whereupon the eye sees the
two images (the specimen and the drawing paper with the pencil) superimposed one on
the other.
Camera lucida appears to have been made for right handed people. In past years
before we were more enlightened, right handed writing and drawing were the rule in
schools, and left handed people had a difficult time.

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 Microscopically Measurement

Mirror

A small hole in the side of

the Camera lucida

Microscope
Drawing paper

Slide

Camera Lucida

How to use a Camera Lucida:

How the Camera lucida is used is that looking through the microscope you see the tip of
the pencil superimposed over the image of the specimen and you draw over the outline
of the image. This, however, is not anything like as easy as it sounds, and can in fact be
difficult and frustrating; some people are able to make perfect drawings of all kinds of
specimens, whilst others find it difficult to use and never master it. Some Victorians
must have mastered its use because many of the drawings of specimens through the
microscope seen in old books were drawn using a Camera lucida.
The microscope to be used need not necessarily be monocular but it should preferably
have an upright eye tube or eye tubes, though only one eye tube is used.
At the side of the microscope place a sheet of drawing paper with a pencil on, lying
beneath the mirror. Temporarily place a small piece of paper on the stage covering the
specimen, so as to block out the image, and bring the Camera lucida top lens into
position and look through. The paper at the side of the microscope with the pencil
resting on should be seen clearly. If not, adjust the swivel mirror at the end of the arm
until the pencil can be seen central on the paper. Remove the paper off the top of the
slide and look through what will be seen as a small bright centre part of the Camera
lucida top lens - it is in this bright centre part that the image appears. There is no need
to refocus the microscope unless you change the magnification. Looking through the

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Camera lucida top lens you should see the image superimposed on the drawing paper
and, in theory, you simply pick up the pencil and draw round the image - if you can at
first see both image and pencil clearly then you are indeed lucky.
Getting the contrast Right:
Usually beneath the Camera lucida top lens is a wheel with a few small supplementary
glasses which are neutral density filters varying in density, the purpose of which is to
vary the relative brilliance as between the specimen and the drawing paper. Looking
through the Camera lucida top lens turn the supplementary filters wheel to each of
these small filters in turn, which may bring up the image of the specimen nice and clear,
but not showing the pencil, or it may show the pencil and paper but no image of the
specimen - you have to find the happy in-between contrast which will show both image
and pencil, though both image and pencil will not now be at their brightest since you are
obtaining the best of both working together. Some Camera lucida have a further one or
perhaps two supplementary filters fastened to the side of the eyecup which can be
swung up into position to give a little more contrast.
Balancing the Light:
Having gone through all that procedure, some people still cannot find the necessary
happy in-between contrast, and here is the secret ingredient - you have to balance the
light intensity as between the illumination of the specimen seen through the microscope
and the light from the table lamp shining on the paper. The microscope illumination is
the prerequisite and is set as standard, it is the light from the table lamp shining on the
paper which has to be controlled and the simplest thing for this is a Woolworths'
standard light dimmer switch fixed between the power point and the lamp.
A few tips:
So, having got set up, looking through the microscope you draw the pencil over the
image seen, but of course the actual drawing you are doing is the size of the magnified
image, i.e., using 5x eyepiece and 10x objective, the drawn image will be about 50
times the size of the image on the slide depending on the position of the drawing paper.
When holding the top lens, particularly when turning the filters wheel; keep your fingers
out of the way of the small hole immediately below and to one side of the Camera
lucida top lens which faces the mirror this is the opening to the tiny reflecting prism in
the Camera lucida head.
The Camera lucida can be used with a microscope having an inclined eye tube, but two
difficulties arise. One is that unless the clamp is securely fixed to the eye tube, the
mirror at the end of the arm swings down with gravity and you have to hold it up. The
other is that the sheet of drawing paper needs to be on a board sloping at the same
angle as the mirror and whilst it is more comfortable to use an inclined eye tube.

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The Graticule:
The graticule, also known an eyepiece micrometer, is a small glass disk etched with a
squared grid for area measurement, or a scale for linear measurement, which can be
fitted in practically any eyepiece.
In the case of the conventional biological microscope, the top of the eyepiece is
unscrewed and the graticule is placed on the field stop, the aperture about one-third to
one-half way down inside the eyepiece which governs the field of view. The field stop is
where the image of the specimen as magnified by the objective comes to rest, and it is
the image at this point which is magnified by the eye lens to give the final image you
see through the microscope. Anything placed at the field stop will also always be in
focus.

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