Apparatus and methods of staining biological slides

MEMORY

Ί

REF: 372,591 & 462,510

SUMMARY OF THE DESCRIPTION

Device for neatly and efficiently coating microscope sample holders with a colored liquid. One or more sample holders are immersed in a liquid bath * that partially fills a chamber in a container. The chamber has a cross section that conforms to the outside of the sample holder so that the immersion of the sample holder raises the level of the liquid and covers all the portions of the specimen holders that a specimen can contain to be colored. To color simultaneously

V a multiplicity of sample holders will these be fixed in a clamping device between the spacer? that they hold parallel and separated a small predetermined distance and the fixed sample carriers are then held by the clamping device during subsequent immersion and extraction.

BACKGROUND OF THE INVENTION Camp; of the invention.

The invention relates in general to the microscopic examination of biological specimens mounted on sample holders. after glass, and specifically includes the coloring of said biological specimens when preparing them for microscopic examination. The coloration and examination of biological specimens takes place daily in a large number of hospital laboratories, emergency rooms, clinics, biology departments, blood banks, and many medical offices, veterinary clinics and hospitals. The sample holders used are glass and generally have a front dimension of 1 x 3 inches (2.54 x 7.62 cm.) And a thickness of 1 mm. Some sample holders have a different front dimension of 25mm x 75mm or a different thickness of 1.2mm. The specimens to be colored and examined can be, for example, tissue, blood sputum or urine. Biological specimens are usually rubbed on the glass sample holder and allowed to air dry, often with heat or reagent fixation. .v, # *, / · '

Previous technique

To color a biological specimen fixed on

J * · · · · a glass sample holder, it is necessary to put a co- or reagent liquid in contact with the specimen in the por-:

ta-samples. It will be noted that for the purposes of the present • · ·

• • description of the term coloration includes the application • · ·>

to the specimen of any of several liquids or re-actives during a coloring procedure, even when the particular liquid does not produce the effective coloration.

In a widely used coloring method of the prior art, the sample holder is placed horizontally, with the specimen above, on a rack suspended in a sump. The dye liquid is poured or dropped drop from a bottle on the sample holder until the specimen is flooded. This technique wastes expensive coloring liquid, because in general a lot more liquid is dropped or dropped than is really needed for coloring. In addition, the sump is colored with surplus liquid and can not be seen after a period of time. The operator's fingers are colored when he picks up the specimen. When the reagents in the bottle remain in the sump area for a prolonged period of time, the fluid may deteriorate or become contaminated, voluntarily or accidentally.

? Q

- 4 _

With this technique, in general, 6 to 8 cm ^ of each reagent are required to carry out a specific examination.

A less common alternative method of the prior art is to vertically place the sample holder or sample holders vertically on a rack, and then to submerge the rack with the sample holder in a colored liquid sample. In this case, too, a much larger amount of colorant is used than actually "•».

for coloring, and, when the reagent is used for more than one sample holder, contamination often occurs. Equally,. This technique involves pouring liquid from bottles into containers and pouring the liquid colorpjaie des., from the containers to the sumps. A third less common procedure still involves using an automatic coloring device. Large hospitals where work is much are more likely to use this method. The machinery necessary for this automatic method is extremely expensive and can only be justified where the coloring task is extremely large. The cost of these machines prohibits their use in most coloring operations. In addition, automatic coloring devices can only be used for one or two colorations.

SUMMARY OF THE PRESENT INVENTION

A specimen holder on which a specimen is fixed is inserted vertically, longitudinally into a closed chamber formed inside a container. Preferably the sample holder is guided and placed inside the camera on rails. A thin film of liquid coloring is formed on each face of the sample holder when

1S _ 5The sample holder moves a bath in the lower part of the chamber to the closed spaces adjacent to the faces of the sample holder. The volume of coloring liquid used is approximately equivalent to the amount of liquid needed to form the thin film on each face of the *

sample holder. Said amount can be only 1 cm.

•> ·

Once closed, the container is resistéorte. manipulation so that the content can not be adulterated before use, thus avoiding a false reading of the colored sample.

When it is removed vertically from the chamber, the colored sample-tube, the coloring liquid is drained, the sample holder and returns to the bathroom in the lower part of the chamber. The container and the liquid used can then be discarded. Can the sample holder be inserted and removed? Take the camera and then operate it by a portion of the sample holder that continuously protrudes from the camera.

Substantially less coloring liquid is required than in the prior art; a fresh batch of reagent is provided for each specimen; the sumps do not receive coloration because the liquid remains inside the chamber; the fingers of the operators do not receive coloration, because the coloring liquid remains inside the chamber; and since the coloration is pre-packaged, the method is faster and more profitable.

The container is a disposable container designed for a single use, and has a lid or shutter that closes the chamber and the liquid until said use takes place. The container is resistant to handling which gives the cer. dfe tea that the coloring procedure will give a true result. Once the coloring is finished, you can again.

- 6 - The camera should be easily eroded before discarding it to avoid any subsequent splash or leakage of camera coloration.

The above embodiment refers to a single sample holder, which also extends to the simultaneous coloration of multiple sample holders.

The idea of ​​the invention is to use .a <>

Thin film of coloring liquid on each face of the specimen holder on which the film is made from a bi. Limited amount of coloring liquid that travels by immersing the sample holder in the container, but this embodiment allows the basic idea to be used with multiple samples.

* * ·. »

The present invention utilizes a clamping device which places, separates and fixes the sample carriers' objects one relative to another. The separation and placement of the sample holders in the fastening device are critical to achieving the effect claimed by the invention, namely, a thin film on the face of each specimen holder that is formed by the sample holders themselves in cooperation with the container when the sample carriers are immersed in a limited bath that occupies only a portion of the container when the sample holders are not submerged therein. The sample holders, while they areseparated, placed and fixed in the clamping device, they are placed in a special coupling vessel containing a limited bath of coloring liquid. The sample holders move the liquid in the bath so that a thin film forms on the faces of the sample holders.

When, with the clamping device, the sample holders are removed from the container, the fluid films

'iΐ' 8 í

%

TO

<img file="ES279935U_D0001.tif" />

The dye falls back into the container and re-forms its substantially the same bath that was originally formed.

The sample holders are then removed from the holding device for further treatment.

Of course, a container, holding device, and co-efficient coloring liquid bath can be produced for a reasonable number of sample holders, but it is not tempting that the most popular use will be the realization of five sample holders. _

BBEVE DESCRIPTION OF TWO DRAWINGS ~

Figure 1 is a perspective view of the sample holder coloring device. . ...

* ·

Fig. 2 is a perspective view of one or more conventional glass samples of metric or English dimensions.

Figure 3 is a perspective view similar to Figure 1 showing the sample holder coloring device with the cover removed and a sample holder inserted.

Figure 4 is a view of a conventional sample holder as shown in Figure 2, which has received coloration in the device of Figure 1.

Figures 5 to 8 are sectional elevational views showing a sample holder in the successive phases of its introduction into the device of the invention shown in Figure 1.

Figure 9 is a graph illustrating the principle involved in the phases of Figures 5 to 8.

Figure 10 is an enlarged sectional elevation view taken on line 10-10 of Figure 1, which shows, in greater detail, the interior design of the device - _{; £}

<img file="ES279935U_D0002.tif" />

<img file="ES279935U_D0003.tif" />

of coloring of figure 1.

Figure 11 is a cross-sectional view taken on line 11-11 of Figure 10, showing other details of the construction.

Fig. 12 is a plan view of the coloring device directly below the lid sheet taken on line 12-12 of Fig. 10, showing the sizes of the entrance opening of the sample holder on the face top of the coloring device.

Figure 13 is a cross-sectional silver view on line 13-13 of Figure 10, showing the profile and proportions of the interior chamber and the r.anbras *, · ^r· «Guide of the sample holder of the coloring device.

»· · ·»

Figure 14 is an enlarged sectional view in cross section a * * * * similar to Figure 10, but showing a sample holder fully inserted into the coloring device, as shown in Figure 3, and showing the combined proportions and the actual positions of the sample holder and the coloring device.

Fig. 15 is a cross-sectional elevational view taken on line 15-15 of Fig. 14, showing the central position of a sample holder supported in slots and the liquid displacement chambers formed on the front faces. and back of the sample holder.

Fig. 16 is a sectional plan view taken on line 16-16 of Fig. 14, showing the relative proportions and positions of the specimen holder and specimen holder staining device, and the liquid displacement chambers formed on the front and rear sides of the sample holder.

• · '-9 - .. ^{: Λ} j - &lt;

Figure 17 is a perspective view of the multi-sample holder coloration device with the sealing cap partially removed.

Figure 18 is an elevational view in section. The tera of the sample holder colouration device 10

<img file="ES279935U_D0004.tif" />

gives on line 27-27 of figure 17.

- ·,

Figure 19 is a sectional elevation view. *.

of the sample holder colouration device taken on line 28-28 of FIG. 17. In FIGS. 2 and 4, a conventional sample holder 20 is shown in detail. The sample carrier 20 is simply a rectangular piece of glass 21 in FIG. which is placed a specimen 22 for microscopic examination. 'Έ1 by * · »ta-samples have opposing flat surfaces or faces'23 and 25, opposite long edges 26 and 27, and short'opposite edges 28.

The most commonly used sample holder has a length (Lg) of about 3 inches (7.62), a width (Wg) of about 1 inch (2.54 cm), and a thickness. (Tg) of approximately 1 mm. In some cases, the sample holder may have an Lg of 75¹¹¹¹¹¹ and Wg of 25 mm or a Tg of 1.2 mm.

These dimensions may vary slightly according to different manufacturers. For example, the thickness can range between 0.0525 inches and 0.0325 inches (0.133 to 0.082 cm), and the width can range between 1.016 inches and 0.964 inches (2.58 to 2.44 cm \

The specimen 22 is prepared and then fixed to one face of the specimen holder in a known manner, for example, using air drying, heat, or reagent fixation.

In general, specimen 22 is placed in the center of one of the sides of the specimen carrier, with a free zone, without specimen, at each end of the face. At this point, the specimen is virtually invisible before coloring.

The sample holder 20, on which the. specimen without coloration 22, is designed to be inserted in a disposable device for discoloration of porta »mu * s *« after 30. It is convenient to make the device 30 of a transparent or translucent plastic in a generally long rectangular configuration having dimensions that they generally conform to the size of the sample holder 20. The device is designed to be used vertically and has an upper part 31, a lower part 32, edges 33 and latios 35 "t.

and 30. A collar or flange 29 surrounds the upper part30 and has a flat surface. ... =

An interior cavity or chamber 40, also of rectangular configuration, is generally adapted to the dimensions of the sample holder 20. The interior cavity 40 has opposed tapered vertical edges 41 and 42, along the vertical edges 43 and 42. 45 of the cavity extending from the top 40 of the cavity to the bottom 47. The cavity also has opposite sides 48 and

The cavity 40 has at its upper end an inclined portion 51 that converges downwards and has beveled edges 52 and sides 53.

A lid or closure 55 is attached to the upper part 31 on the collar or flange 29 with suitable means, such as heat sealing or adhesive. The lid is designed to be removed by grasping and pulling the tongue 56 before use, so that the cavity 40 is exposed.

<img file="ES279935U_D0005.tif" />

. ".eleven"

The cavity contains a bath of coloring liquid 60 that rises up to the middle of the cavity, as shown in FIGS. 1 and 10.

The device is designed to be manufactured in a central station where it closes containing the coloring liquid, as shown in figure 1.

Figures 5 to 8 show sequential views or

introducing a sample holder 20 into the cavity 40.

Figure 5 shows the sample holder 20 on which it is fi xed. a biological specimen 22 to be colored, entering the cavity 40 on the rails 41 and 42. The sample holder 2) is separated from the opposite sides 48 and 49, as shown very well in Figures 15 and 3.6. In Figure 5 the sample holder is about to contact the upper surface of the bath 60, which is the static liquid level. The level of static liquid is at a height H-, above the bottom of the cavity.

In figures 6 and 7 the sample holder progressively descends into the cavity below the bath, the bath fluid moves and rises until the dynamic fluid level reaches H_two, as shown in figure 8, when the sample holder has been fully inserted. At this point, the coloring liquid completely covers the specimen 22. H-jl is approximately half of EL ,. H_two it also represents the height of the cavity, as shown very well in figure 14.

In figure 9 the principle described and shown in figures 5 to 8 is illustrated in graphical form. As the sample holder is submerged below the static liquid level, there is a corresponding increase in the velocity of the sample. dynamic of the liquid coloring virtualmente in relaeiofi of straight line.

When the sample holder is removed from the eolo liquid.

After having remained there for a suitable period of time to carry out the coloring, the sample holder is pulled and the reverse sequence takes place. The level of the liquid drops correspondingly when the liquid is drained from the side of the sample holder to the bath.

Adhere some residual color liquid to the specimen and sample holder and, of course, the height of the ba.

At the end of the coloration, it descends correspondingly below the static level of the liquid, when the sample holder is removed from the device. On the sample holder taken from the JO device is the specimen with color 54, as seen in figure 4. As the device is di. designed for a single use, it is not objectionable this fall in the height and volume of the bathroom.

The tapered or guide portion 51 with beveled edge 52 and the beveled side 53 allows easy introduction of the sample holder into the rails. It also acts as a cavity that collects the coloring liquid that overflows, including the splashes produced when the sample holder falls rapidly into the cavity. TJn collar or guide portion 29 extends around the upper end or upper part of? device and is integral with it. It is convenient that the flange 29 has a flat upper portion 63 that allows to fix ade. therein a sheet or cover 55 with adhesive, or with a suitable heat-sealing technique. The sheet can of being of metal or plastic material that can be removed from the cover by grabbing a portion of tab 56 J pulling^;and&gt;

<img file="ES279935U_D0006.tif" />

from it to the full opening position, as shown in figure 3 · In figures 10 and 11 is the 2 sealing part shown in the cié position? Re and cover.

The width and thickness of the sample holders vary even from those claimed to have specific dimensions. By smoothing the lanes, the₅It will receive a sample holder of varying thickness and width and will continue to keep the sample holder separate from the inside of the cavity. In addition, the tapered lanes ensure that only the edge of the sample holder, and not the surface containing the specimen, touches the lane.

The sample holder 20, once inserted in the: Í and

cavity 40 on the rails 41 and 42 assumes a very defined position j within the cavity and with respect to the sides c walls 48 and 49 of the cavity 40.

As seen in figures 14 to 16 inclusive, the

sample holder 20, once inserted to its lower position j, has an upper portion 61 that comes out of the cavity 40 and the upper part 51 of the device 3 - The upper portion 61 of the sample holder 20 is free of specimen 22, because the specimen zone is conventionally in the center of the sample holder, as shown with lines of transparency in Figure 14. The portion 61 of the sample holder 2^r It is above the device 30, it serves to grasp the sample cutter to insert it into the device and to remove it from the device. The sample holder is also drawn after two of the non-colored dye operator.

grab for this portion during the ena coloring. In this way, those of. they touch a dye solution and are approximately one sixth of the length

<img file="ES279935U_D0007.tif" />

Lg of sample holder protrudes from device 30.

The specimen 22 is colored when the sample holder is fully inserted, as shown in particular in figures 14 to 16 inclusive. As shown in part ^^ Ht in Figures 15 and 16, the specimen holder is placed dettV '; tro of 1? cavity 40 and is separated from the walls or sides 48^one and 49. Beside each of its faces the holder 20, 'has spaces 65 and 66 which contain coloring liquid. The sample holder can be introduced by looking at the specimen 22 in any direction when the sample holder 20 enters the cavity. The face of the sample holder 20 and the specimen 22 on it are positioned so that the specimen 22 can not contact the sides of the cavity, but, on the contrary, the specimen is held so as to be exposed to the specimen. liquid coloring in space 65 or 66 formed; by the sample holder and the side of the cavity. To color the specimen when the specimen carrier is placed like this, a minimum amount of coloring liquid is required. This is shown with clarity in the sectional views of Figures 15 and 16. From the above description it will be seen that, When the sample holder is fully inserted into the cavity, the space 65 and 66 adjacent to each face of the sample holder is a thin flat space containing a suitable amount of coloring liquid to properly color the specimen lying on it, unused excessive liquid. In addition, the sample holder is used to place said liquid next to the faces of the sample holder, and the specimen is on one of the faces. It will be noted that it does not matter which direction the specimen looks when it is inserted into the cavity, because there is a film on both sides of the cavity. In addition, the sample holder is used to place said liquid next to the faces of the sample holder, and the specimen is on one of the faces. It will be noted that it does not matter which direction the specimen looks when it is inserted into the cavity, because there is a film on both sides of the cavity. In addition, the sample holder is used to place said liquid next to the faces of the sample holder, and the specimen is on one of the faces. It will be noted that it does not matter which direction the specimen looks when it is inserted into the cavity, because there is a film on both sides of the cavity.^;sample holder.

The sample holder itself, in cooperation with the container, creates the film together with the faces of the sample holder. It also allows the liquid in said spaces to drain back to the part of the cavity and to the bath formed therein when the sample holder is removed. .

Figures 10 to 16 inclusive show the interior of the device of Figure 1 in detail. W_q is the width of the sample holder 20, Lg is its length.-and Τθ is its thickness. Hg, which is the height of the cavity, as well as the height of the dynamic level of the liquid explained above, is approximately three quarters of the length Lg of the sample holder.

Lanes 41 and 43 include tapered walls at the edges of the cavity, as shown. The width between the rails, W ^, as seen in Figure 13, is slightly larger, for example, 0.02 inches (0.508 mm) greater than the width of the sample holder Wg, so that there is a strike . go suitable for the sample holder to move in the rails. The thickness of the rail at its tapered end, Tqi, is slightly smaller, for example, 0.016 inches (0.406 mm) less than the thickness of the sample holder Tg. The thickness of the cavity, Y, is approximately twice the thickness of the sample holder Tg. By virtue of the tapered rails 41 and 42, the sample holder, once inserted in the rails, is placed in the center between the side walls 48 and 49 of the cavity. When the thickness of the cavity is double that of the sample holder,

IS two on each side of the sample holder, when it is entered -&gt;

The sample holder will be approximately half the thickness. of the sample holder.

Due to the tapered rails 41 and 42, they will be suitably positioned within the sample holder cavity, after having slightly variable dimensions as shown. The tapered walls of lanes 41 and 42 com-

they will think about these slight variations guiding the porta- »· ···· after their edges, placing the sample holder at the same time. Although the sample holder has the smallest width and / or thickness within the variable dimensions, the sample will still be separated from the sides of the cavity, although not all the edges of the holder -Samples can be in firm contact with the lanes. Although said po'rta-mues. After relatively small can have more freedom of movement inside the cavity than a relatively large sample holder, the restriction imposed on the ski, the sample holder by the tapered walls of the rails keeps the sample holder separate from the walls of the cavity so that a film of coloring fluid can be formed next to the faces of the sample holder.

It should be noted that the aforementioned dimensions are merely illustrative and indicative of a cecipient of suitable dimensions for practicing the invention.

To simultaneously display a multiplicity of sample holders, and with particular reference to FIGS. 17, 18 and 19, the specimen carriers are of the type, dimensions and design described above.

The container 130 may be different in

-1710

IS

2S vessel JO described, with regard to shape, height and width, except that the horizontal depth of the container is enlarged to receive a plurality of sample holders, instead of just one, as will be described. . The, . vessel 1J0 has an open top 131, a | lower part 132, edges 133, 134, sides 135 and 13.6, .. and a • »• * • collar or flange 129.

The interior cavity or main chamber 140 of the container 130 has four lanes 141 and 142 in its four vertical chambers, vertical edges 143 and 145, the lower part 147, and opposite sides 148 and 150.

In the version shown, the main camera has • · ..

in its upper part an overflow chamber 160 * shape * jz * * in the flange portion 129 having side walls 168, 170 formed by the leeds of the main chamber 140, terminal walls 172, 174 and lower flanges as 180

A sealing cap 186 is attached to the upper portion 131 of the container, similar to the closure 55 described above.

The sample holders are separated, placed and fixedly fixed in a clamping device.

The container 130 is intended to contain a reduced or limited bath of coloring fluid 360. As can be seen very well in FIGS. 18 and 19, said bath reaches an al. ture H, which preferably reaches a little more than half the height of camera 1 + 0.

As in the embodiment of a single sample holder shown in figures 1 to 16, it is intended that the sample holder 3 be introduced downwards in said bath 360, whereby the coloring liquid of the bath will move upwards in the chamber Mainly substantially has30

18 ..

The upper part of the chamber is not overflowed to the overflow chamber, but in case there is an excessive flow of the displaced liquid, it can be collected in the overflow chamber 160. In the preferred design, the proportion of the parts and the depth of the bath are such that no substantial overflow occurs, and in this case the large overflow chamber can be eliminated and the upper part of the vessel can be made as in the single-sample version of FIGS. 1-6 .

The sample holders are held in spaced relation to each other and to the container so that a thin film forms on both sides of the sample holder. The space between the sample carriers is adapted to the thickness of spacers provided in the clamping device. It is convenient that said thickness is approximately equal to the thickness of the sample holders, which may be approximately 1 mm. The spacing is such that when the spaced sample holders are fixed in the fastening device, and the specimen holders are immersed in the limited bath of the container substantially to the bottom thereof, the bath is displaced so that the coloring liquid rise in thin films that arrive and cover substantially all the portion of the sample holder that penetrates the main chamber of the container and, in particular,

The outer dimensions of the group of five sample holders adapt well to the camera designed to carry out the coloring operations of the sample holders. Said dimensions are such that when the edges of the outer sample holders touch the lanes of the rails 141, 142 in the container, whereby, when the sample holders are introduced into the container, a thin space is formed together to each of the faces i

J outside of the exterior sample holders. The lanes li such as 141, 142 can be integrally molded with the remainder of the container, and to facilitate the insertion of the sample holders they can have entry bevels at their entry ends.

When the sample holders are immersed in the bath, the liquid moves up along the faces of the sample holders. The dimension of the portion of the sample holders that penetrates the holding device is preferably such that when the holding device comes into contact with the upper part of the container, the sample holders almost touch or are slightly above it. the bottom * of the container. In this way, essentially all the coloring liquid is then spread in thin films through the faces of the sample holders and through the specimens placed on the sample holders. The liquid rises to the top of the chamber, and can reach the overflow chamber if one or more sample holders have a slightly larger thickness than the standard sample-plates.

Once the specimens are colored like this, the sample holders are removed from the container by means of the clamping device. The coloring fluid is drained from the sample holders, returning to a bath of the same volume approximately as the initial bath minus the amount that adheres to the sample holder. The container can be closed again with the original closure, and then disposed of neatly.

The above embodiment is illustrated for a group of five sample holders. It should be noted that the same principle can be used with respect to the desired group of, for example, two to ten and more sample holders, each having its container, holding device and bath of specific dimensions. However, it is contemplated that the devices of five sample carriers will be the predominant ones.

As described, the container shown has an overflow compartment. This is generally desirable when the thickness of a bit is expected to vary somewhat. ρο, τ ta-samples used. As the thicker sample holders move more liquid, the overflow chamber serves to receive and contain the overflowing liquid during the coloration operation. When the thickness of the sample holders is relatively uniform, no overflow chamber is needed, because the displacement is more controlled. In this case, the size of the overflow chamber can be reduced, or eliminated as seen in the embodiment shown in Figures 1 to 16.

As an auxiliary means of design, the following relationships are indicated. The distance that the level of the upper part of the dye liquid bath rises when the sample holders are submerged is equal to the volume of liquid displaced upwards by the glass of the sample holders, divided by the average cross-sectional area of ​​the space of the camera without glass to which the liquid moves. Calling at a distance that the sample holders are submerged below the top of the original bath, A ^ to the average cross-sectional area of ​​the chamber by _ 21 _ above the original surface of the bath with the sample holders, after being removed, and A to the average cross-sectional area of ​​the sample-holder glass in the liquid,

H-, ^TOthe/^TOgl ^one

In the simple case of a purely rectangular chamber, five sample holders 1 mm thick and 1 mm apart from each other and from the walls of the chamber, and some submerged sample holders all the depth H of the original bath, the For «mule predicts an increase in the level of liquid

11/5 ~ 1

Ξ = O, 83H.

1,2

In practice, the extra space of the camera at the edges of the sample holders introduced will reduce the height H_two below this value, and this will increase due to the reductions of the space of the camera produced by the rails in the camera.

Although the invention has been described with particular reference to the specific embodiment for perfect clarity, it will be noted that it can be practiced in several ways other than that specifically shown and described, without departing from the spirit and scope of the invention defined by the claims. Attached

In summary, the Utility Model that is requested must fall on the following:

_ 22 "

1S

-