The Microscope and How to Use It

by Dr. Georg Stehli

Overview

A world of pleasure, excitement and new knowledge awaits one who learns to use the microscope — a world in which table salt crystals appear as jewels, a drop of water swarms with life, a butterfly's wings reveal a cascade of multicolored particles. This book is for anyone who would like to enter that world, whether or not he has ever used a microscope before. No special knowledge is required. In non-technical language and with generous use of illustration, the author explains how a microscope works and what kind to use; how to adjust the instrument and position the specimens to be viewed; examination of simple objects: a human hair, feathers, milk. At the same time, he shows how to prepare the objects, what to purchase for the purpose, how to care for it; one's every question is anticipated and clearly answered. The fundamentals understood, the reader is taken into further exploration viewing insect parts, diatoms, plankton, molds, leaves, ferns, fruit rinds, fish scales, animal parts. As we proceed, we learn step by step the techniques involved: use of chloroform, preparation of permanent slides, mounting in glycerine, preparing dye solutions, dissection, and blood smearing. We learn how to detect fat, find Vitamin C in food substances, prepare a frog for examination, view and distinguish bacteria, use the oil-immersion objective, dye bacilli spores, do microphotography, cut sections with the microtome.
Following Dr. Stehli's careful instructions, we have entered and gone well into the fascinating world of microscopy. The invention of the microscope itself started science on new courses, entire fields of new knowledge. The use of a comparatively simple microscope today can start one on a lifetime interest, an absorbing hobby, a career in science, or a permanent addition to one's cultural background. This book provides all the help needed, whether one is adult or student, hobbyist or scientifically serious, seeking education or merely curious about the minute world that exists all about us. 119 photographs and drawings.

Product Details

ISBN-13:	9780486137216
Publisher:	Dover Publications
Publication date:	02/14/2012
Sold by:	Barnes & Noble
Format:	eBook
Pages:	176
File size:	13 MB
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The Microscope and how to use it

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By Georg Stehli, William A. Vorderwinkler

Dover Publications, Inc.

Copyright © 1960 Printed Arts Co., Inc.
All rights reserved.
ISBN: 978-0-486-13721-6

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CHAPTER 1

The Microscope and Essential Tools

A good microscope is the first and most important piece of equipment for your studies. Of course even the simplest and cheapest "magic tube" will unfold many secrets and new wonders; but everyone, once he has got beyond the simplest beginnings, will feel the need to delve further into the apparently puzzling things he sees, and to make more thorough observations requiring greater magnifications—in short, to see more. The disappointment is great when the microscope no longer "co-operates"—when the limits of its powers have been reached.

A really useful microscope is so made that it can be improved with the purchase of additional optical equipment. A good standard model is not very much more expensive than the widely used, but usually disappointingly small, student's microscope. It offers the great advantage that by and by, as your purse permits, it can be made into a complete scientific instrument. A simple and therefore inexpensive instrument is sufficient to begin with, but it should be a model that can be elaborated. For ordinary purposes a in. and a in. objective and a × 5 eyepiece will suffice, but it is advisable to purchase a condenser at the outset. An instrument which is not too expensive and may later be built up is shown in Fig. 1.

HOW A MICROSCOPE IS CONSTRUCTED

The microscope in Fig. 1 rests on a horseshoe-shaped base or foot from which a low, solid column rises. To this column the upper part of the microscope is joined with a simple, quite tight hinge, which enables the body tube of the microscope to be tilted to an angle of 45°; this permits a comfortable position for the head of the observer. A short distance above the hinge is a roomy, square object stage, on which the slide is firmly held by two spring clips. Above the stage there is a sturdy limb which rises above the center of the stage; this holds the tube that carries the lenses. The objective is screwed to the lower end of the tube, and the upper end holds the eyepiece.

For coarse focusing of the microscope there is a rack and pinion adjustment controlled by two large knurled knobs: this moves the tube up and down rapidly. The fine adjustment is made by a second motion controlled by two smaller knobs, the micrometer screws.

With the fine adjustment it is possible to move the tube a minute distance, measured in hundredths of a millimeter. The mechanism for this adjustment is contained within the top of the limb (arm).

The optical arrangement of the microscope is shown schematically in Fig. 2. You see here a cutaway view through the optical axis of an assembled microscope, represented here by line x which passes through the central point of the lenses. C is the system of lenses in the objective. These lenses are compound or achromatic, and give an image that is without colored edges. Each lens does, in fact, consist of two or three separate lenses made of different kinds of glass.

Beneath the objective is the stage t, a metal plate with a hole in the center, on which the slide preparation P is placed and lighted by a small beam. The objective C is screwed into the lower end of a metal tube T which is blackened internally. At the upper end of the tube there is a second system of lenses A. It consists of the two lenses a and d, between which there is usually a diaphragm, and is called the ocular or eyepiece, because it is here that the eye (oculus) of the observer is applied. Under the stage there is an illuminating mirror S, which reflects the necessary light through the hole in the stage on to the slide. It follows that the object being examined must be transparent. As Fig. 2 shows, the optical axis x (the tube axis) passes through the exact center of all parts of the instrument. This is absolutely necessary if the microscope is to deliver sharp, undistorted images.

How the optical components work is shown in Fig. 3. The objective c is represented by only one lens for the sake of simplicity. It projects an exact and therefore photographable image of an object, which is represented by lines a to b. The ocular lens d breaks up these rays which in the diaphragm behind d are united in an enlarged but reversed image b1 to a1. This magnified image is seen through lens e of the eyepiece as if it were seen through an ordinary magnifying glass; the observer sees a repeated, if only weakly magnified, clear image of the object. Because the common Huygenian eyepiece cannot "erect" the image, the view seen in the microscope is reversed. It follows that if you wish to move the examined object to the right or left, or up or down, you must move the slide in the opposite direction to that desired. This may take a little getting used to.

Like any good microscope, the Humboldt may be equipped with several objective lenses (e.g. 1-in., ½-in. and 1/6-in.) and several eyepieces (e.g. × 5 and X 12). The objective lenses are corrected to suit an exact tube length; in the case of the Humboldt this is 170 mm. Objectives and eyepieces can be combined in many ways, so as to give a great range of magnifications (explained in a chart which comes with the instrument). The total magnification of a microscope is determined by a combination of the objective and eyepiece, and is the product of the two magnifications. An objective magnifying 30 diameters, used with an eyepiece of 6 diameters, would give a total magnification of 180 diameters. The magnifying power of the objectives and eyepieces depends upon their focal lengths, and a system magnifies more highly as its focal length decreases.

At this point it might be well to mention that, contrary to common opinion, strength of magnification does not determine the value of a microscope. Much more important than the power of magnification is a microscope's so-called "resolution". This is the power of an optical system to separate minute dots or lines so that details can be distinguished. With a good microscope and the most advantageous lighting the best attainable detail lies around 0.2μ (1μ=1 micron=1/1000 mm.). The tiniest bacteria, which are a little bigger than 0.3μ, can be seen with the help of an exceptional objective—the so-called "oil-immersion lens". Viruses, on the other hand, which are considerably smaller, cannot be seen with an optical microscope at all.

The powers of detail of our eyes are much less than that of a usable microscope—and that is why we can see more with the microscope than with the naked eye. The objective lenses are primarily responsible for the detail rendered, whereas the ocular lenses do not affect detail to any great extent. The objective lenses are therefore the most important (and sensitive) parts of the microscope and must be handled with special care.

Many beginners make the mistake of trying to get immense magnifications by the use of particularly strong eyepieces. It is possible of course to combine a 1/12-in. objective lens with a X 25 eyepiece, for example, and in this way arrive at a magnification of 2,500 diameters. The detail, however, is not increased by the powerful eyepiece, and you cannot see more than with an eyepiece of about X 12. In addition, there is another factor. With unnecessarily powerful eyepieces the image becomes very weakly lighted, the structures no longer appear clear and sharply defined, and there is danger of optical illusions.

The remedy is to find out with which eyepiece you can work effectively with a given objective. The aperture of every objective is designated by a number—either on the objective itself or on a list provided by the manufacturer. The general rule is that the total magnification should never be greater than 1,000 times the number of the aperture. For a 1/12-in. objective (the so-called "oil immersion") with an aperture of 1.30 the highest eyepiece which you can use is X 13. A more powerful eyepiece would lead to "empty magnifications", which can be used only for special purposes, such as the measurement of very small bodies. But there is also a limit at the other end of the scale. The total magnification should not go below 500 times the aperture of the objective, or the detail potentialities would not be fully put to use. This range of magnification between 500 and 1,000 times the aperture is referred to as "usable magnification".

PREPARATION AND AUXILIARY TOOLS

Besides the microscope you will require a little arsenal of preparing instruments, glass equipment and other aids. It is difficult to give the beginner general advice, because the tools necessary for the different branches of microscopy are diversified. Therefore, we will suggest here equipment that will be sufficient only for the very beginning. It is a good idea to keep this equipment all together in a box or "kit":

Standard microscope slides or slips (1 in. X 3 in.). These can be purchased cheaply at any shop where microscopes are sold. The blank slides are traditionally called slips, and those with mounted objects slides, but these terms are becoming interchangeable.

Cover-glasses or coverslips are extremely thin, circular or square pieces of glass for covering objects to be examined. No. 2 or 3, ¾ in. in diameter, are suitable for general purposes, but very thin coverslips are unnecessarily fragile for ordinary work.

Watch-glasses and glass blocks (so-called "salt-shakers") for holding small masses being prepared. You should have 5 watch-glasses about 2 in. in diameter.

Larger stocks of materials, such as bits of twigs, leaves, organs of animals, etc., which are stored in inexpensive preparation-glasses. Old medicinal jars, small preserve jars, etc., will serve the purpose. (Of course you can never have too many glass containers, as you will soon find.)

A thin glass rod about 8 in. long with hemispheral ends, for transferring drops of liquids to slides.

A pair of tweezers made of good steel, for grasping cover-glasses and other objects which would be awkward to handle with the fingers.

A needle-holder with several sized preparation or "setting" needles to tear apart the materials for examination. Ordinary sewing needles may be used in a needle-holder, or permanently fixed into any convenient handle such as a wooden pen-holder. In this way you can make a set of good preparation needles yourself.

Two fine camel's-hair brushes, which are very handy for transferring objects to slides.

A pipette, a glass tube of small diameter, with which small quantities of fluids can be drawn up in order to isolate them for examination. Such pipettes can easily be made. Take a larger glass tube and heat it in the middle in the upper part of a spirit or gas flame, turning it constantly until it becomes red all over. Then pull both ends of the tube apart. Break the thinned-out ends or file them apart. In this way you will get two pipettes, which you can cap with little pieces of rubber for easier handling. This eliminates the necessity of holding your thumb over the end when it is full. An eye-dropper or fountain-pen filler will serve if you are unable to make your own pipette.

To begin with you should get yourself a little wash-bottle (Fig. 5), a spirit lamp with a tripod and wire-netting (Figs. 7 and 8) and some glass bells (Fig. 6), for which purpose preserve jars may also be used. You will need the glass bells to protect open preparations from dust. You can make most of this apparatus yourself. A spirit lamp may be made easily from an ink bottle into which a metal sleeve is fitted to serve as a wick-holder. The tripod can be constructed out of heavy iron wire. The wash-bottle is a little more work. A small flask, perhaps a medicine bottle with a wide mouth, can be converted into a wash-bottle. Take a cork into which you have drilled two holes and insert in the holes two glass tubes, bent over a flame, like those in Fig. 5. If you blow into one tube, the other (which has to be drawn to a fine point) squirts out a thin stream of water. If a greater quantity of water is required, pour it out of the other tube. Wine-glasses, with the stems broken or otherwise, make very good glass bells.

Other tools you will need can be kept in a cigar box padded with cotton-wool. They include:

A scalpel, a pointed knife of good steel, set firmly in a handle. This will serve to cut smaller objects into shape or to dissect larger objects.

A lancet, a needle whose point has the shape of a tiny lance. This is the preferred cutting instrument of the zoologist, because both edges of the point are sharpened. The botanist uses it for transferring larger sections from liquids to slides.

Besides these tools every microscopist needs a sharp pocketknife, a strong pair of scissors and a small pair of embroidery scissors. It is advisable to buy a complete microscopy kit at the beginning, with all the preparation tools in their proper places (Fig. 9).

For chemicals, the basics are: xylol, glycerin jelly and either an artificial resin such as Permount, or Canada balsam, for mounting; methylene benzoate; glycerin; and eau de Javelle (or "Milton") to bleach the preparation. Because the last-named, like tincture of iodine, bites into metal, great care is advised (with the microscope!).

Dye preparations in temporarily usable solution: tincture of carmine, Grenacher's borax, Delafield's hematoxylin, eosin, safranin and Mayer's acid hemalum. Also Löffler's methylene blue.

Outside of these chemicals you will need alcohol. You can do without absolute alcohol, except for fixative purposes. For ordinary examinations, industrial spirit is sufficient, if it is 95%. For closer examinations, to add to dye preparations, to preserve plants and animals, and for dyhydration of objects for permanent preparations, you will use isopropyl alcohol, which is obtainable as 100%, completely free of water. Because it cannot be drunk. it is very much cheaper than absolute alcohol.

You can prepare alcohol of a lower concentration by mixing 95% alcohol with distilled water. Follow these instructions (if a small error is made, it makes no difference) :

Fill into a graduated cylinder as much 95% alcohol as your solution should amount to. (For example. take 50 cubic centimeters [cc.] to get 50% alcohol.) Then fill the graduated cylinder with distilled water to a height of 95 cc. If you have 70% alcohol and want 50%, take 50 cc. and then fill with distilled water to 70 cc.

For most purposes you should keep a supply of 35, 70, 80, 90 and 95% alcohol (for the so-called alcohol stages of dehydration). For 100% alcohol, which you will also need, use water-free isopropyl alcohol. To store the alcohol stages it is best to use 4-oz. or 8-oz. bottles which can be sealed tightly. The liquid should not touch the corks. Alcohol must never be allowed to stand open for any time; it is combustible, evaporates very rapidly, and a high-test alcohol has the unpleasant quality of absorbing water from the air, thereby causing an undesirable dilution.

For dye preparations, you will need acetic acid to be used in an alcohol solution.

Hydrochloric acid, available in 37% solution, will also be needed.

All reagents must be properly labeled. In order not to tip them over, place them in an uncovered cigar-box.

In addition, your work table should have two containers of fresh water for cleaning the preparation tools—a dirty glass rod or a dirty needle could introduce foreign substances into the preparation and spoil everything.

There should be a box of labels to identify permanent preparations.

Elderberry pith and cork should be on hand as well, to hold small objects while cutting them.

A few sheets of filter paper complete the first equipment which the beginner requires for his tasks.

Everyone who works thoroughly keeps records. Your notes should begin when your materials are collected, and should continue through all stages of preparation. It is advisable to keep an individual page for each object, writing in remarks as to the origin of the materials, the manner of preservation and preparation, and also the number of the case in which the preparation is to be placed. Repeat the name of the object in your notebook index.

For these records you can use a bound book, a paper-covered notebook of good writing paper, or a loose-leaf notebook. Those who prefer loose pages may, instead, use index cards (which fit into a box) or cards which fit into a binder.

(Continues...)

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Excerpted from The Microscope and how to use it by Georg Stehli, William A. Vorderwinkler. Copyright © 1960 Printed Arts Co., Inc.. Excerpted by permission of Dover Publications, Inc..
All rights reserved. No part of this excerpt may be reproduced or reprinted without permission in writing from the publisher.
Excerpts are provided by Dial-A-Book Inc. solely for the personal use of visitors to this web site.

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Table of Contents

FOREWARD
1. THE MICROSCOPE AND ESSENTIAL TOOLS
How a Microscope Is Constructed
Preparation and Auxiliary Tools
"Cleaning of Coverslips, Slides and Lenses"
Care of Other Tools Used in Microscopy
2. HOW TO USE THE MICROSCOPE
Adjusting the Prepared Object
How to Get the Best Lighting
Examining and Positioning Preparations
Changing Objectives
Learining to See with One Eye
"Seeing "Flies"
Drawing Microscopic Objects
Fifteen Working Rules for Beginners
3. EXAMINING SIMPLE PREPARTIONS
Table-Salt Crystals
Air Bubbles
Potato Starch
Onion Skin Plasmolysis
Cotton
Nettle Hairs
Canadian Waterweed
Spurge
Cells from Inside the Human Cheek
Milk
Blood
Sperm Cells
Human Hair
Feathers
Cleaning of Slides and Coverslips
4. INSECT PREPARATIONS
Killing with Chloroform
Butterfly Scales
Butterfly Wings
Mouth of June Bug
Eyes of the June Bug
Sting of the Honey Bee
Legs of the Honey Bee
Gizzard of a Cockroach
"Fleas, Bedbugs, Lice and Gnats"
Mites
Storage of Permanent Preparations
5. EXPLORING A DROP OF WATER
Diatoms
Diatom Preparations
Spiorogyra
Mounting in Glycerin or Glycerin Jelly
Water Fleas
Microaquaria and Hay Infusions
Paramecium
Rotifers
Euglena
Amoebae
Fresh-Water Polyp
Preparing Dye Solution
6. THE STRUCTURE OF PLANTS
Moulds
Moss Leaves
Male Fern
Sectioning with a Razor
Care of the Razor
Cornstalks
Cleaning Preparations
Iris Leaf
Orange and Lemon Peel
Nucleus Divisions and Giant Chromosomes
Storage of Fixed Plant Materials
Handling Fresh Sections
Detection of Fat
Detection of Vitamin C
7. THE MICROSCOPIC STRUCTURE OF ANIMALS
Snail Tongues
Liver Flukes
Fish Scales
Fish Blood
Dissection of a Frog
Frog Blood
Ciliated Epithelium of the Frog's Mouth
Striated Muscles
Smooth Muscle Fibers
Columnar Epithelial Cells
Nerve Fibers
Intestinal Infusoria of a Frog
Cartilage
Bone
Bone Section
Pork Liver
Liver Clamps
8. BACTERIA
Dyeing Bacteria
Use of the Oil-Immersion Objective
Nitrogen-Fixing Bacteria
Sulphur Bacteria
Dyeing Bacilli Spores
9. MICROPHOTOGRAPHY
Lighting
Apparatus for Microphotography
Light Filters and Lighting
APPENDIX - MICROTOME TECHNIQUE
Fixing
Dehydration and Paraffin Infiltration
Embedding in Paraffin
Mounting and Trimming the Block
Embedding in Celloidin
The Microtome
Cutting with the Microtome
Subsequent Handling of the Sections
INDEX