Chapter III (2)

_Messrs. Watson and Sons_ have much enhanced their reputation by the marked improvement lately brought about in the manufacture of their whole series of objectives. This probably is chiefly due to the introduction of the _Jena_ glass into their manufacture, and which has enabled them to give increase of aperture to one series in particular, that of the para-chromatic, all of which in consequence are of very high quality. It is difficult to particularise their several objectives, the whole having special features in proportion to their magnifying powers, while much care seems to have been bestowed on them for the elimination of residual colour. A 1/8-inch with correction collar is comprised of a single deep and rather thick front lens, plano-concave flint, and double convex-crown for the middle and triple combination for the back, the latter consisting of two crown lenses cemented to a dense flint (Fig. 124) drawn to scale of 5-1, with lined portions intended to represent the flint, and white the crown glass lenses of the combination. The initial magnification of this objective is 83 diameters, and the numerical aperture ·94. This superior objective can be had for the small sum of £2. Another remarkably useful and cheap objective, their 1-inch numerical aperture 0·21, consists of two achromatic systems forming the front and back with the separation between them of about half an inch, and may also be especially recommended for students’ work.

In the accompanying diagram the lenses are drawn on too large a scale, and therefore the distance between the two combinations should be much greater.

Among the more useful of Watson’s series, the 1-inch, the 1/2-inch, and the 1/6-inch, together with the 1/8-inch dry-objective, and a 1/9-inch, will be found the most serviceable.

_Messrs. Baker_ have their own series of objectives, most of which are so very nearly allied to those of the continental opticians; and what has been said of Zeiss’s and Leitz’s objectives may be taken to apply also to Baker’s, who have an established reputation for their histological series, all of which are well suited for students’ and class-room work.

_Messrs. Swift and Son_ have a new series of objectives, semi-apochromatic and pan-aplanatic, most of which are excellent in quality and show increased flatness of field together with that of achromatism; the index of refraction in each series having been correctly determined together with exact radial focal distance, thus affording more available aperture. I may select for special commendation their 1/12-inch £5 5_s._ homogeneous immersion objective, which is in every way suitable for bacteriological work; its definition is very good, as is seen in a micro-photograph of podura scale, given further on. Their dry 1/6-inch can be had for £1 16_s._--a marvel of cheapness. Of their general series the most useful for histological work are the 1/2-inch, the 1/3-inch at £1 12_s._, and their 1/5-inch of numerical aperture 0·87 at £3.

_Mr. Pillischer_, of Bond Street, has manufactured many excellent objectives. A fine homogeneous oil-immersion 1/12-inch numerical aperture 1·25 is worthy of special notice; it will be found suitable for bacteriological work; it has fine definition with a considerable amount of penetration.

A more intelligent idea of the magnifying power of the objective combined with the eye-piece will be gained by consulting the table given below; precision in this respect has long been a desideratum with microscopists.

Magnifying Powers of Eye-Pieces and Objectives.

A TYPICAL AND INITIAL SELECTION OF POWERS OF EYE-PIECES CALCULATED FOR THE 10-INCH TUBE-LENGTH.

HUYGHENIAN EYE-PIECES.

NAME A B C D E F
OF MAKER. 0 or No. 1 2 3 4 5 6

Baker 6 8 12 15 -- -- Diameters.
Beck, R. & J. 4 8 15 20 25 not made. "
Leitz 5 6 7 8 10 12 " "
Powell & Lealand 5 7·5 10 20 40 " "
Reichert 2·5 3·5 4 5 6·5 " " [24]
Ross 3 8 12-1/2 20 25 40 " [25]
Swift & Son 6 9 12 15 18 21 "
Watson & Sons 4 6 8 10 12 15 "
Zeiss 3 4 5·5 7 9 not made. "

COMPENSATING EYE-PIECES FOR USE WITH APOCHROMATIC OBJECTIVES.

Zeiss 2 4 8 12 18 27 Diameters.

This may be taken as a typical set, further treated of among Eye-pieces.

INITIAL POWERS OF OBJECTIVES CALCULATED FOR THE 10-INCH TUBE-LENGTH.

This is ascertained by dividing the distance of distinct vision 10 inches by the focus of the objective, thus--

Focus-inches 4 3 2 1-1/2 1 2/3 1/2 4/10 1/4 1/5 1/6 1/8 1/12
Initial magnifying
power 2·5 3·3 5 7·5 10 15 20 25 40 50 60 80 120 diameters.

A reference to the above table will at once show that the nomenclature of objectives expresses at once the initial magnifying powers, but as makers have great difficulty in so calculating their formulæ so as to obtain the _exact_ power, these figures must be taken as approximate. Thus a 1/4-inch, which should magnify 40 diameters if true to its description, might actually magnify a little more or less.

The magnifying powers of Zeiss’s and other apochromatic objectives can be ascertained by dividing the focal length of the objective in millimeters into 250 mm. (the distance of distinct vision), thus

Focus millimetres 24 16 12 4 3 2 1·5
Initial magnifying
power 10·5 15·5 21 63 83 125 167 diameters.

The total magnification, when any eye-piece is working in conjunction with an objective, is ascertained by multiplying the initial power of the objective by that of the eye-piece.

The above calculations are all for a 10-inch tube-length. Should, however, a shorter or longer length of body be employed, the magnification can at once be ascertained by a proportion sum. If the magnification be 180 with 10-inch tube-length, what would it be with a 6-inch body--10 : 6 :: 180 = 108 diameters.

Abbe designed three different forms of eye-pieces: 1, the searcher eye-piece; 2, the working eye-piece; and 3, the projecting eye-piece. The _Searcher_ is a negative form of low power. The working is both negative and positive, the positive form of which is constructed on a newer principle; while the projection is chiefly intended for microphotography, its field being small and its definition superlatively sharp. These are severally explained among eye-pieces.

High-Power Objectives.

_Points of Importance for securing the best results with High-power Objectives._--Always give to the body-tube of the microscope the length for which the objective is corrected, 0·160 mm. for the short continental tube, and 0·250 mm. for the English tube (10-inch). Employ both dry and immersion objectives mounted for correction, commencing with a numerical aperture of 0·75 (that is about 100° in air). If the graduation is not given in thickness of cover-glass apply to the maker to correct this omission.

With the homogeneous oil-immersion objective it is highly necessary to utilise all marginal pencils of light, to optically unite the upper lens of the condenser with the preparation as well as the front lens of the objective by means of a liquid having the same index of refraction or at least equal to that of the immersion. _Cedar Oil_ has been generally adopted for the purpose mentioned, the better way of using which is as follows: place a drop on the centre of the front objective, or on the top of the cover-glass, and then lower the objective by means of the coarse adjustment until it comes in contact with the oil, and carefully bring into focus by the fine adjustment. If the slide is held between the finger and thumb of one hand and moved from side to side, while the other hand is working the fine adjustment, there can be no danger of injuring either the objective or the specimen. Before putting the microscope away, take a fine camel-hair brush dipped in ether, alcohol, or methylated spirit, and carefully remove the oil from the objective and the glass cover of the object; a soft chamois leather or cambric pocket handkerchief will dry it off, or a piece of fine white blotting paper answers equally well. Should the lens come accidentally into contact with the Canada balsam, it must be very carefully removed either by ether or alcohol. The former is by far the safest, as alcohol, if not very carefully used, quickly dissolves out the balsam and loosens the cover-glass of the object.

Achromatic Condensers.

_The Achromatic Condenser_ can no longer be classed among the _accessories_ of the microscope, since it is an absolutely indispensable part of its optical arrangements. Its value, then, cannot be overrated, and the corrections of the lenses which enter into the construction of the condenser should be made as perfect as they can be made--in fact, as nearly approaching that of the objective as it is possible to make them. It may therefore be of interest to know something of the rise and progress of the achromatic condenser. In my first chapter I have noticed the earlier attempts made by Dr. Wollaston, whose experiments led him to fit to the underside of the stage of his microscope a short tube, in which a plano-convex lens of about three-quarters of an inch focal length was made to slide up and down (afterwards moved up and down by two knobs); to improve definition he placed a stop between the mirror and the lens. The stop was found to act better when placed between the lens and the object. From this improvement Dr. Wollaston enunciated that “the intensity of illumination will depend upon the diameter of the illuminating lens and the proportion of the image to the perforation, and may be regulated according to the wish of the observer.” Dujardin in France and Tully in England were at work in the same direction. The former a year or two later on contrived an instrument, which he termed an _eclairage_, to remedy the defects of Wollaston’s, and for illuminating objects with achromatic light. This was submitted for approval to Sir David Brewster, who, when the use of the achromatic condenser was first broached, used these encouraging words:--“I have no hesitation in saying that the apparatus for illumination requires to be as perfect as the apparatus for vision, and on this account I would recommend that the illuminating lens should be perfectly free from chromatic and spherical aberration, and that the greatest care be taken to exclude all extraneous light both from the object and eye of the observer.” This far-seeing observer in optical science has borne good fruit, and the outcome of his views is seen in the great development and improvement of the achromatic condenser. In 1839 Andrew Ross made his first useful form of condenser, and gave rules for the illumination of objects in an article written for the “Penny Cyclopædia.” These, epitomised, read as follows: 1. That the illuminating cone should equal the aperture of the objective, and no more. 2. With daylight, a white cloud being in focus, the object has to be placed nearly at the apex of the cone. The object is seen better sometimes above and sometimes below the apex of the cone. 3. With lamplight a bull’s-eye lens is to be used, to parallelise the rays, so that they may be similar to those coming from the white cloud. It has been seen that Mr. Lister foreshadowed the sub-stage condenser.

The early form of Ross’s condenser consists of two small brass tubes made to slide one in the other. To the outer one is attached a flat brass plate which slides underneath the stage of the microscope, and by means of a screw the adjustment of the axis of the illuminator is effected. The upper portion of the apparatus carries the achromatic combination, which by a rack and pinion movement is brought nearer to, or removed further from the object on the stage. The several parts of the illuminator unscrew, so that the lenses may be used either combined for high powers, or separated for low powers.

Messrs. Smith & Beck greatly improved upon Ross’s condenser by adding another achromatic lens to the combination, three being employed when used with high-power objectives and two or even one with the lower, the adjustment and focussing being made by rack and pinion arrangement beneath the stage. Some further changes for the better were made in the condenser by Powell, and in 1850 an amateur microscopist, Mr. Gillett, fully grasping the value of controlling the cone of rays passing into the microscope, devised a new form of condenser, in connection with which a revolving series of diaphragms of different values were made to pass between the achromatic lenses and the source of light.

Andrew Ross constructed the first condenser on Gillett’s principle, and this proved to be one of the most successful pieces of apparatus contrived. _Gillett’s Condenser_ consists of an achromatic lens _c_, about equal to an object-glass of one quarter of an inch focal length, with an aperture of 80°. This lens is screwed into the top of a brass tube, and intersecting which, at an angle of about 25°, is a circular rotating brass plate _a b_, provided with a conical diaphragm, having a series of circular apertures of different sizes _h g_, each of which in succession, as the diaphragm is rotated, proportionally limits the light transmitted through the illuminating lens. The circular plate in which the conical diaphragm is fixed is provided with a spring and catch _e f_, the latter indicating when an aperture is central with the illuminating lens, also the number of the aperture as marked on the graduated circular plate. Three of these apertures have central discs for circularly oblique illumination, allowing only the passage of a hollow cone of light to illuminate the object. The illuminator above described is placed in the secondary stage _i i_, which is situated below the general stage of the microscope, and consists of a cylindrical tube having a rotatory motion, also a rectangular adjustment, which is effected by means of two screws _l m_, one in front, and the other on the left side of its frame. This tube receives and supports all the various illuminating and polarising apparatus, and other auxiliaries.

_Directions for using Gillett’s Condenser._--In the adjustment of the compound body of the microscope for using with Gillett’s illuminator, one or two important points should be observed--first, centricity; and secondly, the fittest compensation of the light to be employed. With regard to the first, place the illuminator in the cylindrical tube, and press upwards the sliding bar _k_ in its place, until checked by the stop; move the microscope body either vertically or inclined for convenient use; and, with the rack and pinion which regulates the sliding bar, bring the illuminating lens to a level with the upper surface of the object-stage; then move the arm which holds the microscope body to the right, until it meets the stop, whereby its central position is attained; adjust the reflecting mirror so as to throw light up the illuminator, and place upon the mirror a piece of clean white paper to obtain a uniform disc of light. Then put on the low eye-piece, and a low power (the half-inch), as more convenient for the mere adjustment of the instrument; place a transparent object on the stage, adjust the microscope-tube, until vision is obtained of the object; then remove the object, and take off the cap of the eye-piece, and in its place fix on the eye-glass called the “centring eye-glass,”[26] which will be found greatly to facilitate the adjustment now under consideration, namely, the centring of the compound body of the microscope with the illuminating apparatus of whatever description. The centring-glass, being thus affixed to the top of the eye-piece, is adjusted by its sliding-tube (without disturbing the microscope-tube) until the images of the diaphragms in the object-glass and centring lens are distinctly seen. The illuminator should now be moved by means of the left-hand screw on the secondary stage while looking through the microscope, to enable the observer to recognize the diaphragm belonging to the illuminator, and by means of the two adjusting screws to place this diaphragm central with the others: thus the first condition, that of centricity, will be accomplished. Remove the white paper from the mirror, and also the centring-glass, and replace the cap on the eye-piece, also the object on the stage, of which distinct vision should then be obtained by the rack and pinion, or fine screw adjustment, should it have become deranged.

The re-publication of the original directions is given with the view of showing what a clear conception Gillett had of the value of his invention. The careful directions given for centring must be regarded with interest, although nearly superseded by the centring screw arrangement in connection with the sub-stage. The best results, he goes on to say, will be secured by using the plain mirror and focussing the window-bar on the object, while a white-cloud illuminator will afford as much light as may be required. It is a mistake to suppose that direct light is more critical than indirect. As a rule, the student is given to over-illuminate the object. These questions will, however, be discussed further on.

Very many modifications of Gillett’s condenser have, since 1850, become known to microscopists. Ross’s present improved form (Fig. 127) is made to drop into the sub-stage of the microscope, and when adjusted, is an extremely efficient instrument. The optical part is similar to a 4/10-inch objective. It has two sets of revolving diaphragms, with apertures and stops for showing surface markings in a perfect manner.

Abbe’s Condenser.

The essential feature of this condenser is its short focus, which collects the light reflected by the mirror, so as to form a cone of rays of very large aperture, having its focus in the plane of the object.

The full aperture of the illuminating cone should only be used when finely granular and deeply stained particles (protoplasm, bacteria, &c.) are being examined with objectives of large aperture. In all cases the cone must be suitably reduced, either by an iris, or other form of diaphragm (_central illumination_). By placing the diaphragm excentrically, by means of rack-work attached to the carrier, the central rays are excluded and a certain extra-axial portion of the illuminating pencil falls upon the object (_oblique illumination_). When the diaphragm is thus excentrically placed, this oblique pencil can be directed from all sides by rotating the carrier round the optic axis. The central stop diaphragm shuts off all the axial and transmits only the marginal rays, thus producing _dark-ground illumination_. The iris diaphragm (Fig. 128) is so shaped that the edge of its smallest opening closely approximates the object-slide on the stage.

The Abbe condenser is the most popular form in use, for all purposes. Owing to the large aperture of the cone of light which it projects, it can be employed with the highest powers; by removing the top lens it can also be used with low powers. Dark ground illumination may be obtained with it up to a 1/4-inch objective.

The condenser is made in two forms of 1·2 and 1·4 numerical aperture by Messrs. Watson. The lenses are mounted in aluminium. Fig. 130 is in more general use, but by workers with high powers Fig. 131 is preferred, as it ensures the most oblique illumination with objectives of largest aperture. It is preferred for photo-micrographic purposes.

_Watson’s Achromatic Condenser_ (Fig. 132), 1·0 numerical aperture, shown in section, although originally designed for use with the micro-spectroscope, is equally efficient for ordinary purposes. This condenser transmits a larger aplanatic cone of light than Abbe’s. It may therefore be employed with higher power objectives, and by removing the top lens it is just as useful a condenser for lower powers. Being constructed with lenses of an unusually large size, it is well adapted for use with the micro-spectroscope. It is certainly one of the best all-round condensers in use. The new Schott glass enters into the construction of the lenses, and these are mounted in aluminium.

Many microscopists consider on the whole that Powell’s sub-stage apochromatic condenser with collar correction (Fig. 133) surpasses that of Abbe. The mechanical arrangement of Powell’s is very simple: the correction collar is similar to that of an ordinary objective, it has a steeper spiral slot and only half a revolution of movement; a long arc is fixed to the collar so that it may conveniently be reached by the finger. It is so constructed as to turn easily and smoothly at the slightest touch. The collar moves only the back lens of the combination, leaving the mount rigid. The diaphragms are regulated by A and B.

The object of the correctional movement is to increase the maximum aplanatic aperture of the condenser by separating the lenses. If the back of a wide-angled objective be examined when an object is illuminated by the full aperture of the condenser, the edge of the flame being in focus, it will be noticed that the illuminated portion of the back lens will be oval and pointed instead of circular. Also that when the condenser is racked up, although the external shape of the illuminated portion becomes more circular, two dark patches will appear on either side of the centre, showing the operation of the spherical aberration of the condenser. If under these circumstances the lenses are separated by means of the collar adjustment, the black spots will be closed up, and a circular and evenly-illuminated disc of illumination of a larger size will result. The wheel of diaphragms, or a series of graduated diaphragm discs to drop into a holder, is intended for critical work; the diaphragm can always be recorded, and the identical illuminating cone reproduced.

Hence we have a simple method of graduating apertures between any two contiguous diaphragms; if, for example, we place the lever to the left, so that the lens may be separated as far as possible, and use a No. 6 diaphragm, and if, on examining the object, it is thought that the illuminating cone is not large enough, and if when No. 7 is turned on it is found too much, we can go back to No. 6, and by turning the lever 60° towards the right, closing the lenses and increasing the power a little, we shall obtain an aperture somewhere between Nos. 6 and 7 diaphragm. Thus we can by means of the correction collar graduate the aperture with the facility as with an iris, and we can record any particular aperture with a degree of accuracy foreign to the iris. It must be admitted, however, that the cone of light transmitted by the condenser is a very small one.

Powell also supplies an apochromatic oil-immersion condenser, numerical aperture 1·40, but without collar correction; Fig. 134 shows the sliding tube lowered by arm A and cell B withdrawn for changing stops, which can be done without altering the focus of the condenser. Fig. 134_a_ shows the cell B closed and raised by arm A close to the back lens of optical combination. In Fig. 134_b_ six of the principal stops are shown. Powell’s dry apochromatic condenser, of nearly 0·9 aplanatic cone, is also very good; but the high price of all is a bar to their more general use. The speciality of these is the conversion of axis light into condensed oblique incident light by the refraction of the condenser.

Messrs. R. & J. Beck have various forms of achromatic condensers, some of which partake of a somewhat elaborate arrangement; others are simple and inexpensive, to suit the students’ microscope; as when the light of the concave mirror proves insufficient for any object requiring intense transmitted light, an achromatic condenser must be adapted to even the students’ form of microscope. The latest form of condenser (Fig. 135) is fitted with revolving stops and iris diaphragm, and other appliances for obtaining satisfactory results.

_Beck’s Compound Illuminating Apparatus_ (Fig. 136).--It is useful in working with the microscope to be enabled to rapidly change the illumination, and for this reason this compound form of condenser has been constructed. It consists of an upper portion A, a wide-angle condenser, the aperture of which can be reduced at will by an iris diaphragm, moved by the lever B. This can be used for all other purposes. Below this diaphragm is a plate C, which can be swung back out of position at will, as shown in outline. Into a cell in this plate the stops D can be dropped, and the condenser can be used for dark field illumination, or for high powers as an oblique illuminator. A large-size polarising prism E, fastens to the plate C, and can be removed when not required. In this way any of the various modes of illumination may be separately or conjointly obtained.

Their condenser (Fig. 137) has a large aperture, and facilities for rotating the series of diaphragms. It is available for either dry or immersion objectives up to 1·3 numerical aperture on diatoms, and wet or dry histological objects. The spherical form of the front is worked by a milled-head that rotates a series of lenses and diaphragms. It also avoids the inconvenience of having the connecting fluid drawn away by capillary attraction, as would be the case if mounted on a flat surface. It is also less in the way of the sub-stage movements.

_The Parachromatic Condenser_ of Messrs. Watson (Fig. 138) was made to meet a demand for a condenser giving a large solid cone of illumination free from colour. The optical part of this condenser consists of a full hemispherical front lens, and the middle and back combinations of such forms as to produce the necessary corrections. The Jena phosphate crown and silicate flints are used in its manufacture, and to these are due its special qualities. The total aperture of the condenser is 1·0, and it yields an aplanatic aperture of ·90 numerical aperture. The magnifying power is 2/7ths of an inch. From this it will be seen that it is especially suitable for use with high-power objectives.

It can also be employed without the front lens, when the magnifying power is 4/10ths of an inch, and the numerical aperture ·35. It is mounted in an exceedingly convenient manner, the iris diaphragm being fitted in such a way as to be absolutely central with the optical system.

The arc through which the handle controlling the iris travels is divided, and indicates the aperture at which the condenser may be working at any time. An important feature in this condenser is that it is almost wholly free from colour. As a rule condensers of the same form are found difficult to work with, because of the small diameter of the field or back lens. This difficulty has been successfully overcome by increasing the size of this lens, and the whole of which is fully utilised.

Most London opticians have their own especial form of achromatic condenser, designed for and fitted to their several stands and objectives, varying from a small price to the more expensively-fitted accessories.

Messrs. Swift’s illuminating apparatus (Fig. 139) is conveniently supplied with numerous useful appliances. The optical combination A is computed to be used as an effective spot lens from a 3-inch objective up to a sixth. C C are two small milled heads by means of which the optical combination A is centred to the axis of the objective. The revolving diaphragm E has four apertures for the purpose of receiving central stops, oblique light discs, and selenite films. D is a frame carrying two revolving cells, into one of which a mica film is placed, which can be revolved with ease over either of the selenites below, whereby changes of colour can be obtained in experimenting with polarised light. The darts and P A’s indicate the position of the positive axis of the mica and selenite films, and by this means results can be recorded, etc. Either of the revolving cells can be thrown into the centre of the condenser, and there stopped by means of a spring catch; when so arranged the mica film, &c., may be revolved in its place by turning the cell D, as both cells are geared together with fine racked teeth. F is a polarising prism mounted on an eccentric arm, rendered central when in use, or thrown out, as seen, when out of use. G is the rack dove-tail slide for indicating and focussing the condenser on the object. The advantages associated with this condenser consist in having the polarising prism, selenite films, dark-ground, and oblique light stops, so that they may be brought close under the optical combination.

Baker’s Nelson Condenser, shown in Fig. 140, is intended for use with their medium instruments. It has, however, many pieces of apparatus essential to those of a higher class. It is applicable, indeed, to all instruments having sufficient depth beneath the stage to receive it. It comprises an achromatic combination of 90° aperture, available with all powers up to 1/8-inch tinted glass for neutralising the yellow rays of artificial light, focussing adjustment, dark-ground illuminator, large diaphragm with rotating tube to carry oblique light stops, small wheel of apertures, polarising prism with two selenite films, clear aperture, and oblique light-shutter for low powers.

Baker’s Students’ Condenser (Fig. 141) is designed to take the place of Abbe’s, and costs much less. It transmits a larger aplanatic cone of light, and can be used either with high or low powers by removing the front lens. It is equally useful for photo-micrographic work.

Mr. J. Mayall’s semi-cylinder or prism for oblique illumination (Fig. 142) is a convenient form, as it permits of the semi-cylinder being tilted and placed excentrically; in this manner, without immersion contact, and by suitable adjustment, a dry object can be viewed with any colour of monochromatic light. If placed in immersion contact with the slide, the utmost obliquity of incident light can be obtained. Objects in fluid may be placed on the plane-surface of the semi-cylinder, and illuminated by ordinary transmitted light, or rendered “self-luminous” in a dark field, as with the hemispherical illuminator or Wenham’s immersion paraboloid. A concave mirror with a double arm is quite sufficient to direct the illuminating pencil. This semi-cylinder was originally made by Tolles, of Boston, for measuring apertures, but, at Mr. Mayall’s suggestion, Messrs. Ross mounted it as an illuminator.

The spiral slot should be fixed close beneath the larger lens of the condenser, and when properly arranged will be found a convenient mode of obtaining oblique light.

_The Webster-Collins Universal Condenser_ (Fig. 143) is so well known that it scarcely calls for any lengthy description. It is an inexpensive form of condenser, designed in the first instance for use with the students’ microscope. It is fitted into the sub-stage; has an iris diaphragm as well as a series of revolving diaphragms moved by a milled head screw arrangement.

Oblique Illumination.

_Wenham’s Parabolic Condenser._--Mr. Wenham’s many useful additions to the microscope and its accessories demand especial notice. When mention is made of the various immersion condensers (illuminators, as he preferred to call them), his original right-angled prism, his truncated hemispherical lens, his immersion paraboloid, and his reflex illuminator, in which rays beyond the angle of total reflexion are utilised by reflex action from cover-glass on to the surface of the object, every one of these well-devised inventions will always be spoken of in terms of praise. All in their turn conferred a great service upon the microscope, and enabled the student to clear up difficulties that stood in the way of developing structure when achromatic lenses and dry-objectives were considered perfect. The superior illumination of the object was wholly due to, and effected by, _reflected_ rays from the object to the aperture of the objective, and obviously, reflex action could only take place with dry-objectives. This reflex action must be regarded as Mr. Wenham’s special discovery. It must be observed, however, that it is not the same as the more modern achromatic appliances used for throwing _direct_ rays upon the object, and which proved the existence of apertures capable of direct transmission up to 27° measured in the body of the front lens.

The most practical of Mr. Wenham’s inventions is probably the hemispherical lens, since adopted by Messrs. Ross in connection with their excellent Zentmayer stand, and which has proved eminently serviceable. But the fact is that devices of the kind for obtaining direct oblique light require a thin stage, and therefore most of those who possess the earlier-made microscope stand would doubtless hail the appearance of any appliance which will convert axial light into oblique light; as by so doing the possessors of such instruments, in which the stage is generally of considerable thickness, would enjoy the pleasure of seeing the best resolution it is possible to get with their dry-objectives.[27]

_Wenham’s Parabolic Reflector._--This will be better understood by reference to Fig. 145, which represents it in section A B C, and shows that the rays of light _r r′ r′′_, entering perpendicularly at its surface C, and then reflected by its parabolic surface A B to a focus at F, can form no part of the largest pencil of light admitted by the object-glass and represented by G F H; but an object placed at F will interrupt the rays and be strongly illuminated. A stop at S prevents any light from passing through direct from the mirror.

In the microscope the _parabolic reflector_ fits into the cylindrical fitting under the stage, and the adjustment of its focus upon the object is made by giving it a spiral motion when fitted in--that is, carefully pushing it up or down at the same time that it is turned round by the milled edge B B. It must then be focussed by the rack and pinion motion. As the rays of light must be parallel when they enter it, a _flat mirror_, which in this case should be added to the instrument, is generally used; daylight will then require only direct reflection, but the rays from an artificial source will have to be made parallel by placing a side condenser between the light and the mirror, about 1-3/4 inch from the former and 4-1/2 inches from the latter. Nearly the whole surface of the mirror should be equally illuminated; this may be tested by temporarily placing upon it a card or piece of white paper. Parallel rays can also be obtained from the concave mirror, if the light is placed about 2-1/2 inches from it. Dark-ground illumination is not suitable for very transparent objects--that is, unless there is a considerable difference in their index of refraction, or they are pervaded by air-cells.

One very remarkable example of this may be seen in the tracheal system of insects. If any of the transparent larvæ of the various kinds of gnat be mounted in gelatine and glycerine jelly, slightly warmed but not enough to kill the insect outright, about the third day the fluids circulating in the body will be absorbed and replaced by air. Illuminated by the parabolic condenser, and viewed with a binocular microscope, and a low power, the gnat-larva becomes a superb object. The body of the insect is but faintly visible, and in its place is displayed a marvellous tracheal skeleton, with the tubes standing out in perspective, shining brilliantly, like a structure of burnished silver. Unfortunately, such objects are not permanent, for when the whole of the water dries up, the tracheal tubes either collapse or become refilled with fluid.

As to the blackness of field, and luminosity of the object, this depends upon excess of light from the paraboloid received beyond the angle of aperture of the object-glass. It is found in practice that more and more of the inner annulus of rays from the paraboloid has to be stopped off, until at last, with high-angled objectives, it is scarcely possible to obtain a black field.

The light, on the whole, most suitable for this method of illumination is lamp, the rays of which should in all cases be rendered more parallel by means of a large plano-convex lens, or condenser.

_Wenham’s Immersion Condenser._--Mr. Wenham, in the year 1856, described various forms of oblique illuminators, one of which was an immersion; a simple right-angled prism, connected by a fluid medium of oil of cloves. This, however, was abandoned for a nearly hemispherical lens connected with the slide, and although an improvement, did not touch the point of excellence Mr. Wenham was looking for. Ultimately he adopted a semi-circular disc of glass of the exact form and size represented in the drawing, Fig. 146, having a quarter-inch radius, with a well-polished rounded edge, the sides being grasped by a simple kind of open clip attached to the sub-stage, the fluid medium used for connecting the upper surface with the slide being either water, glycerine, or oil; an increase of oblique illumination being obtained by swinging the ordinary mirror sideways. By means of an illuminator of the kind difficult objects mounted in balsam are resolved. This simple piece of glass collects and concentrates light in a marvellous manner, and is by no means a bad substitute for some of the more costly forms of achromatic condenser. It can be used either in fluid contact with the slide, or dry, as an ordinary condenser.

Mr. Wenham subsequently contrived a small truncated glass paraboloid, for use in fluid contact with the slide; water, glycerine, oil, or other substance being employed as a contact medium. The rays of light in this illuminator, being internally reflected from a convex surface of glass, impinge obliquely on the under surface of the slide, and are transmitted by the fluid uniting medium, and internally reflected from the upper surface of the cover-glass to the objective. To use the reflex illuminator efficiently it must be racked up to a level with the stage. The centre of rotation is then set true by a dot on the fitting, seen with a low power, a drop of water is then placed on the top, and upon this the slide is laid. Minute objects _on the slide_ must be found either by the aid of a low power, by their greater brilliancy, or by rotating the illuminator; the effect on the podura scale is superb, the whole scale appearing dotted with bright blue spots in a zig-zag direction. Objects for this illuminator should be especially selected and mounted.

The Amici Prism, originally designed for oblique illumination, consists of a flattened triangular glass prism, the two narrower sides of which are slightly convex, while the third or broadest side forms the reflecting surface. When properly used, it is capable of transmitting a very oblique pencil of light. The prism is either mounted, as in Fig. 147, for slipping into the fitting of the sub-stage, or on an independent stand, as arranged for Powell’s microscope, page 85, Fig. 56.

Method of Employing the Achromatic Condenser to the Greatest Advantage.

_Its Illumination._--Good daylight is the best for general work. The microscope should be placed near a window with a northern aspect. Direct sunlight should never be utilised; the best light is that reflected from a white cloud. A good paraffin lamp is the most serviceable artificial source of light, and it is quite under control. As an illuminant more often brought into requisition in the smoky atmosphere of towns, the paraffin lamp is on the whole the handiest and the most useful. If gas-light can be brought into use as suggested for micro-photography, with the incandescent mantle, it will be found to be the purest and best form of artificial illumination for the microscope. Among paraffin lamps those constructed by Baker and Swift are all that can be desired.[28]

Sectional view of the Optical Arrangement of the Aplanatic Bull’s-eye Lens, fitted in gymbal on the front of the lamp.]

As the chimneys of these lamps are made of metal, and blackened, no reflected light disturbs the eye. Care must be taken to have the wick evenly trimmed; the metal chimney has a glazed front, giving exit to the rays of light, the flat of the flame being used with low powers, and the image of the flame being reflected by a plane mirror to give equal illumination of the whole field. In working with high powers, the lamp is turned with the flame edge-wise, and at the same time the mirror must be dispensed with. By working, as it is termed, directly on the edge of the flame, the illumination is greatly increased, and a band of light can be concentrated on any part of the preparation it is desired to make a careful study of.

To obtain the best results, time and care must be given to the illumination of the object. The lamp and microscope having been placed in position, a low power is first used and the smallest diaphragm. On looking through the microscope it will probably be observed that the image of the diaphragm is not in the centre of the field; by moving the centring screw of the condenser this may be adjusted. The low power is then replaced by a high power, the largest diaphragm used, and the bacteria or diatom brought into focus. The diaphragm must now be replaced by one of medium size, and by racking the condenser up and down, a point will be arrived at when the image of the edge of the flame appears as an intensely bright band of light. If this is not exactly in the centre of the field the centring screw of the condenser must again be adjusted. With regard to the use of diaphragms, various sizes should be tried while focussing with the fine adjustment, at the same time using the correction colour; in this way we obtain the sharpest possible image. When the condenser has been accurately centred, it will still be necessary to focus it for each individual specimen, so as to correct for difference in the thickness of slides and the layers of mounting medium. Correction for different thickness of cover-glasses must be made by the aid of the collar adjustment in the following way: a high-power eye-piece is substituted for the ordinary eye-piece, and the faults in the image will thereby be intensified. By moving the collar completely round, first in one direction and then in the other, while carefully observing the effect of the image, it will be seen to become obviously worse whichever way the collar is turned. The collar must then be turned through gradually diminishing distances until an intermediate point is reached at which the best image results with the high-power eye-piece, and on replacing this by the low-power eye-piece the sharpest possible image will be obtained.

_Effect of the Sub-stage Condenser._--The sub-stage condenser gives the most powerful illumination when it has been racked up until it almost touches the specimen. It produces a cone of rays of very short focus, and the apex of the cone should correspond with the particular bacterium or group of bacterias under observation. The effect of the condenser without a diaphragm is to obliterate what Koch has termed a _structure picture_. If the component parts of a tissue section were colourless and of the same refractive power as the medium in which the section is mounted, nothing would be visible under the microscope. As, however, the cells and their nuclei and the tissues do not differ in this respect, the rays which pass through them are diffracted, and an image of lines and shadows is developed. If in such a tissue there were minute coloured objects, and if it were possible to mount the tissue in a medium of exactly the same refractive power, the tissue being then invisible, the detection of the coloured objects would be much facilitated. This is exactly what is required in dealing with bacteria which has been stained with aniline dyes, and the desired result can be obtained by the use of the sub-stage condenser.

If we use the full aperture of the condenser the greatly converged rays play on the component parts of the tissue, light enters from all sides, the shadows disappear, and the structure picture is lost. If now a diaphragm is inserted, so that we are practically only dealing with parallel rays, the structure picture reappears. As the diaphragm is gradually increased in size the structure picture gradually becomes less and less distinct, while the colour picture, the image of the stained bacteria, becomes more and more intense. When, therefore, bacteria in the living condition and unstained tissues are examined, a diaphragm must be used, and when the attention is to be concentrated upon the stained bacteria in a section or in a cover-glass preparation the diaphragm must be removed and the field flooded with light--(Crookshank).

The wide-angle condenser, it will be understood, consists of a combination of lenses, which concentrate all the light entering them to a small point, and the condenser must be so accurately focussed that this brilliant cone of light, when it emerges from the upper lens of the condenser, falls upon the object from all directions, forming a wide-angle cone of light, at the apex of which the object must be placed (see Fig. 149). That is to say, the object is illuminated by a cone of rays passing through it in all directions.

There are, however, objects which require a fully illuminated field, when the lamp should be turned round and the Herschel lens condenser (shown in section, Fig. 148) should be used to collect the light and throw it upon the mirror. For moderate powers, as a four-tenth or one-fifth, the condenser should be used a little below the focus to give an even illumination over the whole field. Moreover, as to the use of the condenser for defining general objects, it must be borne in mind that to show different kinds of structure different apertures in the iris diaphragm are necessary, and that whereas some objects show their structure better with a large angle of light cut down in intensity by the use of blue glass, others show better with a small pencil of direct rays. For the resolution of diatoms it is often necessary to use oblique light only, and for this purpose diaphragms with central patches are used, the iris diaphragm being opened to its full extent. An annular ring of oblique light emerges from the condenser upon the object, and it is in this manner also that dark-ground illumination is obtained with moderate and low powers.

THE DIAPHRAGM.

The early form of diaphragm in use was that shown in Fig. 150.

It consists simply of a circular brass plate with a series of circular openings of different sizes, arranged to revolve upon another plate by a central pin or axis, the last being also provided with an opening as large as the largest in the diaphragm-plate, and corresponding in situation to the axis of the microscope body. The holes in the diaphragm-plate are centred and retained in place by a bent spring in the second plate, which rubs against the edge of the diaphragm-plate and catches in a notch. The blank space shuts off the light from the mirror when condensed light is about to be used. It is usually made to fit in under the stage of the microscope. This has been almost superseded by the iris diaphragm, originally designed by Wales, of America. It was made by this optician for his working students’ microscope. An early form of the iris diaphragm is seen in Fig. 151. By pressing upon the lever handle at the side the aperture gradually closes up, and without for a moment losing sight of the object under examination.

The Mirror.

The mode in which an object is illuminated is, in the words of the late Andrew Ross, “second only in importance to the excellence of the glass through which it is seen.” To ensure good illumination the mirror should be in direct co-ordination with the objective and eye-piece; it must be regarded as a part of the same system, and tending by a combined series of acts to a perfect result. Illumination of the object is recognised as of three kinds or qualities--reflected, transmitted, and refracted light. For the illumination of transparent objects, transmitted light is brought into use; for opaque objects, reflected light is needed.

The mirror should be about 2-1/2 or 3 inches in diameter, and it must not be fixed, but made to slide up and down the stem under the stage, so that the rays of light emanating from it may be brought to a focus. The utility of the mirror is so obvious that it is occasionally passed over in silence by writers. To myself it appears to be an important accessory of the microscope, and I shall therefore proceed to combine theory with practice in what I have to say with regard to the mirror.

The microscope mirror should be the segment of a true sphere, and its centre that of a true curvature. If the mirror has a true circular boundary, the central point on line A (Fig. 152) of the reflecting surface, is the pole of the same. The line A C is known as its principal axis, and any other straight line through C, which meets the mirror, is its secondary axis. When the incident axis is perfectly parallel to the principal axis, the reflected rays converge to a point F, its principal focus. So much for the theory of the mirror. Now we come to its practical use.

Simple as the mirror of the microscope may appear to be, if the curve of the surface is not perfect, it will yield a secondary reflection or double pencil of rays. The plane mirror will occasionally be found to emit more than one reflection of the lamp-flame; this we find may be corrected by rotating the mirror in its cell. Many years ago I proposed to meet a difficulty of the kind by arranging a rectangular prism on a separate stand, shown in Fig. 153, consisting of a prism A B, mounted in gimbal C, D, and E, secured to a brass tube G, fitted to the stem, and thus made to take the place of the mirror.

The direct method of employing the mirror, that more generally resorted to, is by reflecting rays from the concave surface; the plane surface is preferred when the condenser is used. Whichever is employed, it should not be forgotten that the _optic axis_ must be preserved throughout, and so brought to the centre of the open tube of the microscope. Another method is to interpose a bull’s-eye lens, and in this way supply the mirror with a beam of parallel rays of light. The plane side of the bull’s-eye lens should be turned towards the lamp, so that lamp, bull’s-eye, sub-stage condenser, and objective, are brought into an exact line, the bull’s-eye being set at right-angles to the line. A piece of thin white paper held across the bottom of the sub-stage will serve to show whether the rays of light are fairly parallel. The next care is to focus the object on the stage, and then the sub-stage condenser on the slide; further correction should be made by means of the centring screws of the sub-stage, or by moving the bull’s-eye lens or lamp slightly, thus perfecting the arrangements for working with parallel rays of light.

Accessories of the Microscope.

The accessories and appliances of the microscope have become so very numerous, that any attempt to describe them and explain the uses to which they are put would demand more space than I find myself in a position to bestow upon them. I must therefore confine my remarks to those accessories in more general use.

Having described the method of employing transmitted light, I have a few words to add with regard to the illumination of opaque objects by reflected light. A very early and efficient form of opaque illumination is the well-known _Lieberkühn_. This has not been entirely surpassed by more recent inventions. The concave speculum termed a Lieberkühn, so named after its celebrated inventor, directly reflects down upon the object the light received either from the mirror or bull’s-eye lens. It consists of a silver cap, which slides over the objective (Fig. 154), _a_ indicating the lower part of the compound body, and _b_ the objective over which slides the Lieberkühn, _c_; the rays of light are collected to a focus upon the object at _d_. The object may either be mounted on a slip of glass, or held by the stage-forceps, _f_; if very small, or transparent, it may be gummed to the dark well, _e_, or mounted on a Beck’s opaque disc-revolver.

This holder will be found useful for the examination of opaque or other objects that cannot be conveniently held by the stage forceps, the specimen being temporarily attached to it by gum or gold size. The holder is intended to rotate, so that every portion of the object can be brought into view. In this way it will be found useful in the study of insects, foraminifera, &c.

With the Lieberkühn, however, the illumination of opaque objects must be more or less one-sided, and therefore, the silver side-reflector has superseded it for general use (Fig. 157). To ensure a more perfect illumination of the object, the bull’s-eye lens should also be used. Mr. Sorby devised a reflector to fit over the objective. It consists of a semi-circular cap; is, in short, a modification of the parabolic reflector. The light from the mirror can, by slightly varying its inclination, be brought into use with this reflector.