US2244082A - Apparatus for and method of maintaining and working with biological specimens in a germfree controlled environment - Google Patents

Description

June 3, 1941. REYNlERs 2,244,082 APPARATUS FOR AND METHOD OF MAINTAINING AND WORKING WITH BIOLOGICAL SPECIMENS IN A GERMFREE CONTROLLED ENVIRONMENT Filed June 14, 193"! 7 Sheets-Sheet 1 r4; m w //%/07 m: w

l 47 INVENTOR u BY mgm

4/- JAMES A Eff/W565.-

June 3, '1941. Y J. A. REYNIERS 2,244,082

APPARATUS FOR AND METHOD OF MAINTAINING AND WORKING WITH BIOLOGICAL SPECIMENS IN A GERMFREE CONTROLLED ENVIRONMENT Filed June 14, 195': 7 Sheets-Sheet 2 INVENTOR. JAMFJ A. Eff/W565.

June 3, 1941. REYNIERS- 2,244,082 APPARATUS FOR AND METHOD OF MAINTAINING AND WORKING WITH BIOLOGI AL ASPEC IMENS IN A GERMFREE CONTROLLED ENVIRONMENT Filed June 14, 1937 7 Sheets-Sheet 4 I 7 5 INVENT RY J MEJ ,4.'-.,:r/y/5eJ-- AT OR EY;

June 3, 1941. REYNIERS 2,244,082 APPARATUS FOR AND METHOD OF MAINTAINING AND WORKING WITH BIOLOGICAL SPECIMENS IN A GERMFREE CONTROLLED ENVIRONMENT Filed June 14, 1937 7 Sheets-Sheet 5 INVENTOR. JAMfiJ 4. Firm/52.5

BY mamm ATTORNEYS.

June 3, 1941. J A, Y s 2,244,082 APPARATUS FOR AND METHOD OF MAINTAINING AND WORKING WITH BIOLOGICAL SPECIMENS IN A GERMFREE CONTROLLED ENVIRONMENT.

Filed June 14, 193'! 7 Sheets-Sheet 6 INVENTOR' JAMFJ ,4 Eff/W525.

ATTORNEY 5 June 3, 1941- ,J. A. REYNIERS 2,244,082 APPARATUS FOR AND METHOD OF MAINTAINING AND WORKING WITH BIOLOGICAL SPECIMENS IN A GERMFREE CONTROLLED ENVIRONMENT Filed June 14, 1937 7 Sheets-Sheet 7 ATTORNEY-5.

Patented June 3, 1941 UNITED STATES PATENT OFFICE APPARATUS FOR AND METHOD OF MAIN- TAINING AND WORKING WITH BIOLOGI- CAL SPECIMENS IN A GERMFBEE CON- TBOLLED ENVIRONMENT James A. Reyniers, South Bend, Ind. Application June 14, 1937, Serial No. 148,102

20 Claims.

This invention relates to apparatus for, and method of maintaining and working with, biological specimens in a germ-free,- controlled environment. More particularly, this invention re-' lates to method and means for operating, examining, raising, maintaining, storing, handling, and breeding animals or plants in a germ-free and controlled environment without risk of contamination.

Various methods and means have heretofore been developed and attempted for the purpose of obtaining, maintaining and working with biological specimens in a germ-free condition and controlled environment for biological and medical research and experimentation; but such have universally been found to be inadequate in yarious essential respects. The greatest difficulty which has been heretofo're experienced with methods and means for this purpose has been the inability to maintain the specimens under the required germ-free and controlled conditions for a length of time adequate to conduct the desired research or experiments; or to permit the preparation, reproduction and growth of the specimen as is necessary for long range tests. Thus, such prior methods and means have failed to provide means for introducing fresh sterilized food and water to the specimens, and to accommodate transfer of the specimens from one container to another while maintaining the requiredgerm-free controlled environment at all times preparatory to, during, and after the transfer. Where the specimens are animals, the need from transfer from time to time to permit cleaning of the cage or other container is obvious, as is also the need for replenishing the food and water supplies and for changing the character of the food as the animal passes through the various phases of a life cycle.

It is, therefore, the primary object of this invention to provide a method and means for maintaining biological specimens in a sterile or germ-free condition and controlled environment for any desired period of time, and to maintain successive generations of animals in such condition and environment.

A further object of this invention is to provide a method and means for working with biologica1 specimens while in a germ-free controlled environment without risk of contamination of the specimens.

A further object is to provide a method and means for transferring biological specimens from one container to another while maintaining its germ-free, non-contaminated condition and the desired controlled environment or atmospheric condition.

A further object is to provide a method and means by which food, water and supplie may be introduced into the apparatus and su plied to the specimen whenever desired or necessary without exposing the specimen to contamination or altering the environment or atmospheric condition in which the specimen is maintained.

A further object is to provide a plurality of containers constructedto accommodate particular phases of the treatment or examination of a specimen and which are readily connectible in hermetically sealed relation, with means for sterilizing a selected container and means for selectively sealing connected containers from each other, said last named means being controllable without danger of contaminating the sterilized containers.

A further object is to provide an apparatus of this character comprising a hermetically sealed, sterilized container, with means for affording access to the interior thereof for manipulation thereof and of the specimens contained therein without contaminating the same.

A further object is to provide an apparatus of this character which may be sterilized while a specimen is contained therein without injuring the specimen.

A further object is to provide a device of this character with novel means for sterilizing the same while desired matter, such as micro-organisms, are contained therein.

A further object is to provide a steam-sterilized apparatus of this character with means for insulating selected connected portions containing specimens against the heat of the steam.

A further object is to provide a sterilized or germ-free apparatus of this character with means for maintaining selected temperature and humidity conditions therein without contamination.

A further object is to provide apparatus of this character by which bacterial and chemical tests, or tests to determine immunology, may be conducted under germ-free conditions.

A further object is to provide apparatus of this character into which material may be inserted, or from which material may be removed, without exposing the specimen contained in the apparatus to contamination.

Other objects will be apparent from the description and the appended claims.

In the drawings:

Fig. 1 is a side view illustrating one arrangement or assembly of the apparatus, and including a raising, an operating and an examining unit interconnected for conjoint use in manipulating and treating the specimen.

Fig. 2 is a side view illustrating another arrangement or assembly of the apparatus, including a raising, a storage and a transfer unit interconnected for conjoint use.

Fig. 3 is a side view illustrating another arrangement embodying two storage units and an view taken on line 99 of Fig. 5.

Fig. 10 is a fragmentary transverse sectional view similar to Fig. 9 and illustrating a modified construction provided with a water jacket.

Fig. 11 is a side view illustrating a unit having an access or connecting member in operative relation thereto.

Fig. 12 is a fragmentary vertical longitudinal sectional view of the arrangement illustrated in Fig. 11. r

Fig. 13 is a fragmentary longitudinal sectional view illustrating the construction and arrangement of a connector between two units of the device.

Fig. 14 is a side view of one of the units with the outer doors of the glove clave in open position.

Fig. 15 is a transverse sectional view of a unit taken on line I5I5 of Fig. 14.

Fig. 16 is a fragmentary enlarged longitudinal sectional view. taken on line I6-I6 of Fig. 15 and illustrating the glove clave construction.

Fig. 17 is a fragmentary transverse sectional view taken on line I'|-I'| of Fig. 16 and illustrating a portion of the glove clave.

Fig. 18 is a fragmentary sectional view of the glove clave illustrating a modified construction of the glove mounting.

Fig. 19 is an end view of the glove clave closed position.

Fig. 20 is an interior end view of theglove clave.

Fig. 21 is a transverse sectional view of' the glove clave taken on line 2I2| of Fig. 1'1.

Fig. 22 is a fragmentary longitudinal sectional view illustrating a storage unit connected with another unit of the apparatus.

Fig. 23 is a side view of a storage unit mounted on a support and independent of the remainder of the apparatus.

Fig. 24 is an end view of the storage unit and its support.

Fig. 25 is a fragmentary enlarged longitudinal sectional view illustrating the means for interconnecting the storage unit with another unit sectional view taken on line 28-28 of Fi 29. Fig. 29 is a fragmentary vertical transverse sectional view taken on line 2928 of Fig. 28. Fig. 30 is an enlarged fragmentary longitudinal sectional view of the lap cloth of the operating unit.

Referring to the drawings; whichillustrate the preferred embodiment of the invention, and particularly to Figs. 1 to 4, the numeral I designates a raising unit, 2 an operating unit, 3 an examining unit, 4 a transfer unit, 5 a storage unit, and 6 a connecting or access unit. Each of these units will be more particularly described hereinafter, as will the particular means for interconnecting and manipulating the same for conjoint use as required.

Each of the following: raising unit I, operating unit 2, examining unit 3, and transfer-unit 4, are essentially of the same basic construction, and are all desirably of the same size and especially of the same diameter. The construction of these units is best illustrated in Figs. 5 to 10. Essentially each of these unitscomprises a sheet metal cylinder I0 whose longitudinal seam (not shown) is suitably sealed, as by welding. To each end of cylinder Ill is secured an end ring II (Fig. 7) of generally angle section. Adjacent to the inner portion of the ring II in the cylindrical flange thereof is formed an annular groove I2 in which the end of cylinder I0 snugly fits, the outer portion I3 of said grooved flange extending beyond the inner portion thereof, whereby the same may be secured together by, rivets I4. The joint between the portion I3 and the cylinder wall may be continuously welded or otherwise treated to form a seal. The radially outwardly projecting flange of .the ring is provided with spaced apertures I5. The outer face of ring II, preferably opposite groove I2, is provided with an annular groove in inwardly spaced relation to apertures I5 to receive an annular gasket I6 of any suitable sealing material, said gasket normally projecting beyond the outer face of ring II as illustrated in Fig. 7.

In the center of the cylinder III at its uppermost portion is formed an opening II, preferably substantially circular. A ring I8 whose lower surface conforms to the contour of cylinder III at the margin of opening I], and preferably having an outwardly projecting flange- I9 conforming to the cylinder contour, is secured to said cylinder 'by means of rivets 20 passing through flange I9 and into the cylinder. The joint between ring I8 and cylinder I0 is sealed, as by gaskets or welding. The upper edge of ring I8 is slightly annularly recessed at its inner margin to receive an annular gasket 2|. A plurality of spaced longitudinal screw threaded bores are formed in ring I8 at the upper end thereof in outwardly spaced relation to gasket 2|. A circular plate 22 of heavy glass which can withstand the application of a high degree of heat and steam such as "Pyrex," is supported at its margin by gasket 2|, the diameter of plate 22 being substantially the same as the outer diameter of said gasket. An annular gasket'23 similar to gasket 2| is superimposed on the margin of plate 22. A retaining ring 24 of angle section fits over and around said gasket 23 and plate 22, and is provided with a pluralityof apertures registering with the bores in the end of ring I8 to receive bolts 25 which securely fasten ring 24 to ring I8 and compress gaskets 2| and 23 to form a seal. By this construction a positively sealed sight opening is provided in the cylindrical body of each unit.

One or more pairs of holes 26 are formed in cylinder II in complementary relation and adjacent end rings These holes 26 are preferably iisposed with their centers approximately 30 degrees above the horizontal center plane of the :ylinder, as illustrated in Fig. 6. A ring 21, best illustrated in Fig. 9, having a plane outer edge and an inner edge conforming to the contour of the cylinder is secured to. the cylinder at the .nargin of each hole or opening 26. These rings 21 are preferably provided with spaced longiudinally extending screw threaded bores at their opposite ends. Bolts or screws 28 passing through he cylinder and into the bores at the inner ends f said rings 21 serve to secure the same to the ylinder. Suitable means, such as gasket rings :11 welded joints, provide the necessary seal etween said rings 21 and the cylinder.

In instances where the rings at the openings 26 are to be provided with cooling means, a ring construction as illustrated in Fig. 10 is employed. In this construction a ring 36 of U-section is provided with a lower or inner flange 3| shaped to conform to, and fit in face contact on, cylinder l8. -Rivets 32 or other suitable means serve to secure the flange 3| to the cylinder; and suitable means are provided to seal the joint between said flange and cylinder. The upper or outer flange 33 of ring 30 is plane. Annular flanges 34 project from the inner faces of both ring flanges 3| and 33 in opposed relation, and an annular groove is formed in flange 33 with its inner wall forming a continuation of the outer wallof the adjacent flange 34. A strip of sheet metal 35 fits into said groove at its outer or upper edge and engages the outer faces of opposed flanges 34 to which it is secured by rivets 36. The seam of said strip 35 (not shown) is welded or otherwise sealed whereby an annular water chamber is formed at the exterior of ring 30 defined by flanges 3| and 33 and strip 35. One or more holes may be formed in strip 35 to accommodate passage therethrough of the end portions of water tubes 31 which are encircled by suitable gaskets 38 mounted in the openings in strip 35. Suitable spaced apertures 39 are formed in the outer portion of outer flange 33 and radially outwardly spaced from strip 35.

Each cylinder I is preferably mounted upon a suitable standard 40 here illustrated as comprising legs 4| secured at their upper ends to the cylinder adjacent the ends and at the under side thereof. These legs are interconnected and reinforced at spaced points by cross bars 42. These standards are of the same height for each of the individual units built upon a cylinder Ill.

The openings 26 afford access to the interior of the units whereby the operator of the apparatus may reach into the various units for handling and manipulating the specimen, food, instruments or the like as necessary for the treatment, maintenance, examination or other required technique. For this purpose rubber gloves of a length sufllcient to extend from the openings 26 to and preferably beyond the ends of the unit are employed. It is essential that these gloves be so mounted that the surfaces thereof which are exposed to the specimens are absolutely germfree, and that they are so sealed in their mounting that the use thereof cannot contaminate the apparatus or the specimen. An illustrative form of glove mounting of this character is illustrated in Figs. 14 to 18. In this construction a glove clave 45 is mounted on the water jacketed ring 33, though it will be understood that the same may also be applied to the plain ring 21 iilustrated in Fig. 9. ,The glove clave 45 comprises an outer cylindrical member 46, an inner cylindrical portion 41 spaced from portion 46, and end walls 48 integral with and closing the space between the inner and outer cylindric l portions to provide an annular water jacket 49. The outer end wall 46 is inwardly spaced from the outer end of outer cylindrical portion 46, and is preferably of stepped outer formation best illustrated in Figs. 16 and 18, for purposes to be hereinafter set forth. The ends of suitable water supply and discharge conduits 56 may be threaded in the outer portion 46 to circulate water in water jacket 49. Walls 5| integral with, and spanning the space between, inner and outer portions 46, 41 are provided in the clave and preferably extend diagonally-relative to the clave axis. These walls 5| each have a passage 52 extending longitudinally. therethrough for communication with the clave interior, and terminate at their outer ends in enlarged threaded portions 53 for connection with steam lines 54. A flange 55 projects radially outwardly from the clave intermediate its ends and is connected by suitable bolts 56 to flange 33 of ring 30, said bolts passing through suitable apertures in flange 55 in registration with apertures 39 in flange 33. An annular gasket 51 is preferably interposed between flanges 33 and 55. The outer diameter of clave 45 is preferably sufficiently less than the inner diameter of ring 38 to facilitate easy assembly of the parts.

The stepped form of the outer end wall 48 ac-.

commodates a ring 58 of angle section secured thereto adjacent its outer margin by bolts 59 threaded into suitable tapped bores in said wall 58. The outer end ,of a glove 66, preferably formed of rubber, fits between said end wall 46 and ring 58 to be locked to place thereby when said ring is tightly applied, said glove at the same time serving to form a seal when tightly pressed between said ring and wall to prevent access of air or dirt into the interior of the clave. In Fig. 18 is illustrated a modified form of glove mounting wherein the outer end of glove 60 is fitted around a ring 6| seated in an annular inner marginal recess in end wall 48 and pressed tightly against said end wall in sealing relation by a flat ring 62 secured by suitable bolts 59 to wall 48 in the manner previously described.

An annular flange 63 projects interlorly from inner clave wall 41 at the inner end of the glove clave, and is provided with a pair of opposed cutaway portions 64. Against this projecting flange 63 the ends of a cross bar 65 are adapted to bear. Bar 65 has a central threaded aperture through which extends a threaded shaft 66 mounting an operating handle 61 at its outer end. Shaft 66 is journaled in the center of an inner closure plate 68 having an annular rib 69 projecting therefrom and adapted to seat against an annular gasket 10 mounted in a suitable groove in the inner end wall 48. An arm 1| is integrally formed with the 'outer clave wall adjacent its inner end and projects inwardly therefrom. To the end of arm 1| a link 12 is pivoted at 13, the other end of said link being pivoted at 14 to part 15 projecting from plate 68 and accommodating rotation of the shaft 66 with which it is connected. It will be seen that, when the ends of bar 65 bear against flange 63 and handle 61 is rotated to cause shaft 66 to draw closure plate 68 toward said bar, the rib 69 will seat firmly against gasket 19 to seal the interior of cylinder l6 from said clave 45. To open the closure plate, shaft 66 is rotated to loosen the engagement of bar 85 with flange 03 and permit said bar to be turned on said shaft into registry with cut-outs 84 to there.- by relzease the plate for swinging movement on link I An arm 16 is formed integrally with and projects forwardly beyond the end of outer clave wall 48, and this arm pivotally mounts at its outer end a bar 11 extending diametrically across the outer end of the clave. Bar 11 is provided with a central threaded aperture in which a threaded shaft 18 is mounted. The outer end of shaft I4 carries a handle", and a head 80 is formed at its inner end. Head 80 flts in a recess formed centrally in the outer face of a closure plate 8L.

and a retaining plate 82secured to plate 8I and encircling shaft 18 serves to connect said shaft and plate. An annular groove is formed in the inner face of plate 8| adjacent its margin,- and receives a gasket 83 which is adapted to bear against th outer edge of the outer cylinder 40 of the clave. Opposite arm I8 is provided a projecting lug 84 to which is pivoted a link 85 against which a hook portion 86 on the end of bar 11 is adapted to bear. It will be seen that when hook 86 engages link 85 and shaft I8 is rotated to urge bar 11 outwardly, the closure plate is inwardly urged to tightly seal the outer end of the clave 45, and a reverse rotation of said shaft will permit disengagement of hook 88 from link 85 to permit pivoting of said closure plate 8| to open position.

In the manipulation, treatment and maintenance of the specimen it is necessary that means be provided for the purpose of transferring the specimen from one chamber or unit to another while maintaining the specimen-in a germ-free controlled environment at all times. The construction by which this purpose is achieved is best illustrated in Figs. 11 to 13, and involves the use of the connecting or access member 6. As here shown, member 6 is provided with an outer cylindrical wall 90, an inwardly spaced concentric cylindrical wall 9 I, and integral annular end walls 92 interconnecting said cylindrical walls and providing a closed annular water chamber 93 between the cylindrical walls. The outer wall 80 preferably projects beyond the end wall 92 at each end of the device. Arms 94 project from the member 5 at each end thereof and to the ends of these arms are pivoted at 95 the cross bars 98.

hooked end portions '91 which are adapted to be held by links 88 pivoted at 99 to ears I projecting from the member 6 in opposed relation to the complementary arm 94. Each bar 96 is provided with a threaded opening through its center for the reception of a threaded shaft IOI mounting a handle I02 at its outer end and connected to a closureplate I03 at its inner end for rotation relative to said closure plate. The closure plate I08 is provided with an annular recess in its inner face which-mounts an annular gasket v I04 adapted to be pressed against the edge of the projecting end of outer wall 90. A plurality of plugs I05 are formed integrally with and extend between the inner and outer walls 90 and 8| and have threaded passages I08 therethrough for connection with conduits I01 which form a steam supply system. Threaded apertures I08 are formed in the outer wall 90 for connection with conduits I09 which form a water circulating system.

Each of the connectors I5 is provided with one or two radially outwardly projecting flanges H0 adjacent its ends, for connection with the adjacent unit or units of the apparatus. One form of connection between unit 8 and the adjacent units is shown in Fig. 12 wherein an adapter ring III is connected by bolts II2 to end ring II of the adjacent unit and projects radially inwardly therefrom. A cylindrical flange portion III integral with ring III at its inner margin and preferably outwardly spaced from outer wall 00 of the connector, carries a radially outwardly projecting flange H4 spaced from and parallel to ring III. The adapter is thus of channel section, and is provided with opposed annular flanges II5 on the adjacent faces of portions III The opposed ends of bars 06 are provided with mens.

and I I4. A strip of sheet metal I I6 is tightly secured at its margins to said flanges I I5 to form an annular water jacket. An annular gasket II! is interposed between adapter flange H4 and connector flange I I0, and securing members I I8 connect said flanges. In practice, the adapter rings III, II4 are connected in operative relation to the connector unit 6 before the same is secured to the adjacent unit and its end ring II for convenience in handling and assembly.

A modified construction for mounting unit 8 is illustrated in Fig. 13, wherein a plane adapter ring I20 is connected to end ring II of the adjacent unit to which unit 6 is to be connected. In this form, the inner marginal portion I2I of the adapter is preferably of greater thickness to permit the formation therein of tapped bores which extend only partially therethrough, and which receive the inner ends of bolts I22 passing through flange IIO of the connector 6 to secure the adapter to the connector unit. In radially inwardly spaced relation to said bolt holes the adapter ring I20 is provided with an annular groove in its outer face to receive an annular gasket I23 against which flange IIO seats to form a positive seal between rings H0 and I20.

It will be seen from the above that the units are so arranged that specimens may be readily .passed from one to another thereof, the whole apparatus being positively and hermetically sealed to maintain the desired germ-free or controlled environment therein during the transfer. Likewise, when necessary or desirable, each of the connected units may be positively sealed from the remaining units of the assembled apparatus, as when a specimen is contained in one unit and food, equipment or other material is to be introduced into the apparatus and then steam sterilized before exposure of the specimen thereto. It will, of course, be understood that the provision of the sealed glove claves in the various units, as heretofore described, is incorporated in the various large interconnected units to permit opening and closing of the various closures between the units and to permit transfer of the specimens, food, etc. from one to another of the connected units.

The storage units 5 afford means by which one or more specimens may be maintained in the desired controlled environment while theremalnder of the apparatus is being independently used for the treatment, maintenance, etc. of other specimens. In other words, the storage units 5 may be employed to store individual or groups of specimens while the remainder of the apparatus is used for treating and manipulating the speci- An illustrative construction of the storage chamber is best illustrated in Figs. 22 to 25 wherein chamber 5 comprises a slightly tapered circular wall'd25 and an integral annular end wall portion I26 providing a central circular opening I21 at the small end of the chamber. Opening I21 is provided with a sealed window construction wherein a plate I28 of heat resisting transparent material is held between opposed complementary rings I29 of angle section which mount gasket rings I30 to effect a seal between the same and plate I23. Rings I29 are drawn tightly together and connected to the wall I26 by securing members I3I radially outwardly spaced from transparent plate I28, a gasket ring I32 being interposed between the inner ring I29 and said wall, with said securing members preferably passing therethrough.

At the opposite or large end of the unit, the wall I25 is provided with an integral outwardly projecting circumferential flange I33 preferably spaced slightly from the edge of said wall. The outer periphery of said flange preferably includes a tapered portion I34 best illustrated in Fig. 25. A ring I35 flts closely around wall I25 adjacent flange I33, a gasket ring I36 being interposed between said ring and flange, and a plurality of longitudinally extending securing bolts I31 connecting said ring and flange and drawing ring I35 tight against said gasket. Ring I35 is provided with a tapered portion I38 in its outer periphery in opposed relation to portion I34 of flange I33, and is also provided with a portion of its inner periphery tapered at I39 in complementary or parallel relation to outer tapered portion I38.

For the purpose of connecting the storage unit to one of the other units of the apparatus, an adapter ring I40 is secured to the end ring II of said unit. Ring I40 is provided with a substantially cylindrical inner marginal flange I4I adapted to fit closely on Wall I25, and said flange terminates in an outwardly tapering flange portion I42. A gasket ring I43 is interposed between flange I42 and ring I35 to be pressed against the wall I25 by the inner face of flange I 42 and the tapered inner face I39 of said ring and thereby eifect a seal between unit 5 and adapter ring I40. Suitable securing means, such as clips I44 bearing upon the opposed tapered outer faces of flange I42 and ring I35 serve to hold said parts together and tight against jacket I43 to effect the necessary seal.

The unit 5 is provided with a closure plate I45 at its open enlarged end, the periphery of said plate including a portion tapered in opposed relation to face I34 of flange I 33 whereby a plurality of securing members I46 similar to members I44 may be applied thereto to hold said closure plate in sealed relation on said unit. A seal is effected between wall I25 and plate I45 by providing an annular groove in the inner face of said plate, which groove mounts a gasket I41 if desired and into which the free forward edge of wall I25 closely fits.

Unit 5 is provided with a suitable flat supporting plate I48 at its lower side to support the specimen, and brackets I49 are associated therewith to mount suitable receptacles (not shown) for holding food and Water for the specimen. Adjacent the upper end of the unit is provided an air filter I50 mounted on a suitable holder I5I having nipples I52 for connection with conduits (not shown) for controlling the circulation of air, thus afiording means for controlling the temperature and humidity of the air supplied to the unit. It will be seen that, after a specimen has been transferred into unit 5, a supply of food and water provided therein, and the closure plate I45 applied in sealed relation thereto, the storage unit 5 may be removed from the apparatus. To achieve this purpose the adapter I40 is removed from the adjacent unit to disconnect the units, and securing members I44 are then released to permit separation of the adapter from the storage unit. A suitable support may be provided to hold the storage unit independently of the remainder of the apparatussaid support comprising a frame I53 including arcuate cross pieces, and legs I54 mounting said frame. The air fllter I50 permits disconnection of the storage chamber from a controlled air supplied source during the time the unit 5 is being manipulatedand transferred to and from the apparatus and a storage place having a separate air connection, if such is desired.

Where the specimens to be treated and maintained in the apparatus are animals, such as guinea pigs, it is essential that the same be in a germ-free, non-contaminated condition when inserted in the apparatus. For this purpose, I

employan operating unit 2 best illustrated in Figs. 26 to 30. This unit is provided with ring I8 at its upper center for mounting the transparent plate 22 as illustrated in Fig. 8; and preferably with two pairs of glove claves 45, as illustrated in Fig. 16, on opposite sides of the body or cylinder I0 to permit two operators to work together. The unit is supported upon the usual supporting structure 40, but the upper portion of said structure is preferably enclosed to form a trap. The trap is formed by side and end walls I 55 which extend around the frame and flt tightly, and preferably in sealed relation, to the cylinder I0 at their upper edges. A bottom with openings through which the legs 4| extend in a tight sealed fit completes the trap. Panels I56 formed of transparent material are mounted in the side and end walls, and an access door I51 is provided in one of the walls.

An opening I58 is formed in the center of cylinder I0 at the lowermost portion thereof, said opening preferably being of rectangular projected outline. A frame I 59 having an upper .marginal flange I60 fits tightly in face engagement with the outer surface of cylinder I0 at the margin of opening I58, said frame being secured to said cylinder by securing members I6I and preferably sealed at said joint. A pair of integral depending arms I62 project from frame I59 at one side thereof and extend downwardly below said frame. To the lower ends of these arms I62 are pivoted at I 63 links I64 which are connected at I65 to brackets I66 mounted on a closure plate I61. A groove extends around frame I59 at its lower edge and mounts a gasket I68 against which the margin of said closure plate bears. Suitable securing means, such as bolts I69 passing through the plate I61 outwardly of gasket I68 and threaded into frame I59 serve to connect said plate in sealing relation to said frame. A pedestal or support I10 is carried centrally by the upper face of plate I61, and mounts an operating plate I1I. An inner frame I12 fits tightly against and is secured to marginal posi tion of cylinder I0 around opening I58. A lap cloth is carried by inner frame I12 and extends across the opening I58 in cylinder I0. The lap cloth is clamped to place at its margin in sealed relation by a rectangular frame I13. The lap cloth is best illustrated in Fig. 30, and comprises two thicknesses of cloth or other suitable flexible material I14 which are provided with registering central slits I15; and between these cloth layers is interposed a sheet I16 of suitable material,

such as "Cellophane" which can withstand application of steam to maintain a seal, and which is normally imperforate. The inner frame I12. also serves as a support to which suitable plates Ill may be secured in substantially flush relation pointof entry of said leads therein.

The raising unit I and transfer unit 4 are each of substantially the construction heretofore described as built around cylinder III as a body and provided with standards l0, sealed sight openings, sealed glove claves and other features as hereinbefore described. The transfer unit 4 being employed merely to facilitate transfer of the specimens from one to another of the other units, usually contains no additional equipment. The raising unit I, examining unit 3 and such other units as may be required are of the same general construction above described, with the exception that they may be provided with suitable equipment, such as electric leads III for lighting and other purposes, and in the event of the examining unit I, with a microscope IOI. mounted in sealed relation to the cylindrical body of the unit and with a grinding or pulverizing machine I82 mounted on a suitable support I on the unit and having its operatin portion (not shown) extending into the cylindrical body with a sealed connection.

The method of use of the apparatus is as follows: Assuming that it is desired to initiate use of the apparatus for animals, such as guinea pigs, selected units will be interconnected and sterilized. Thus an operating cage 2, and a raising cage I, may be interconnected by a connecting unit 8, the same constituting a hermetically sealed assembly which can be sterilized by the introduction of steam and then cooled to desired temperature by the water jacket as above described. A pregnant animal, preferably denuded of hair and bathed in an antiseptic solution to insure germ-free condition as far as external or skin conditions are concerned, is placed in the trap of the operating unit through door I51. The animal is then held on operating table III in desired position while closure I81 is raised and locked in operative position. The lap cloth Ill-I16 forms a primary hermetic seal between the cylindrical body of the operating unit and the trap while the animal is being placed on the operating table, and thereafter, upon lockingof closure plate It! in place, said plate forms a secondary hermetic seal. It will be understood, of course, that the trap of the operating unit is cleaned, as by an antiseptic solution, preparatory to placing of the animal therein. When the animal is in proper position and closure I6! is sealed, the intermediate layer I16 of the lap cloth may be slit complementary to slits I15 in outer layers I14 whereby access may be had to the animal. A Caesarian operation may then be performed on the animal to remove the young, as by use of an electric cauter or knife, precautions being taken during the operation as by antiseptic treatment of exposedorgans of the animal, to prevent entry of germs into the unit. The newly born animals may then be removed through access or connecting unit ,6 to raising chamber I containing the necessary equipment for handling, treating and feeding the young after birth. The closures I03 (Fig. 12 of connecting unit I may be sealed after the young are in the raising cage I, so that operating cage 2 may be disconnected therefrom if desired.

It will be understood that each of the raising and operating units contains the sight opening construction illustrated in Fig. 8 by means of which the operator may see the animal and all other contents of said units during these operations. Likewise it will be understood that these units are provided with the glove clave construction illustrated in Figs. 14 to 18, so that the operator may, by inserting his hancb and arms into the rubber gloves 60, reach into the respective Interior condition of these units. The rubber gloves are flexible, and of a length suflicient to permit the operator to also reach into the connecting unit 6, as to pass the animals therethrough from one primary unit to another and to operate the handles I02 controlling the closures III of said connecting unit.

Assuming now that the animal has reached a stage where it can feed itself, and it is desired to transfer the same to a storage unit 8, a hookup as illustrated in Fig. 2 may be made, wherein there are connected-to the assembly of raising unit I and connecting unit I, a transfer unit 4 at the end of connecting unit 8, and a storage,

unit 5 at the end of said transfer unit. During the time the connection of these additional units is mechanically eiiected by the use of the interconnecting members illustrated inl igs. 13 and 22, at least one of the closures III of the connecting unit 0 is maintained in sealed relation. After the mechanical interconnection of the units is effected, steam or other sterilizing media, as antiseptic solution, is used to thoroughly sterilize the newly connected units. When this sterilizing operation is completed, the closures Ill of connecting unit i may be opened and the animal passed successively through said connecting unit 6, transfer unit 4, and into storage unit 5. It will be understood that the food and water supply is inserted in the storage chamber 5 before sterilization thereof where steam is the sterilizing medium, so that the same is subjected to the sterilizing action and. is insured to be in germ-free condition. After the animal is inserted in storage unit 5, the closure I45 thereof may be applied to seal the same from the remainder of the connected apparatus, whereupon the parts or units may be disconnected for storage of unit 5 in a suitable place having connections for air filter I50. During such storage time the unit 5 may be supported on standard I 53-454 (Fig. 24).

It will be seen that the animals can be held in any one storage unit 5 for only a limited period of time, as determined by food and water supply or other conditions, and that the animals must, therefore, be transferred to another storage unit from time to time. For this purpose an assembly as illustrated in Fig. 3 may be employed, comprising a transfer unit 4 mounting storage units 5 at each end thereof. The unit 5 whichcontains the animal will be maintained in sealed condition while the assembly of parts is effected, and while the empty units are being sterilized. Thereafter the units may be opened into communication by the use of the glove claves to open the respective closures and then handle the animal, as to transfer it to the freshly conditioned storage unit.

It will be obvious from the above that the various units may be assembled in any desired groups or systems to permit transfer of the animals from one to another unit, as from storage to examining cage, without at any time exposing the animal or other specimen to atmosphere or to any contaminating influence. Likewise, the specimen may be maintained in the desired germfree condition for any desired period of time which may range from a short time to a full life span or even to successive generations. In the latter connection, it will be obvious that where the specimens are animals, they may be brought together for breeding purposes or the like, in which event the new generation may be born in the natural way without necessity for resort to the Caesarian operation technique employing operatlng unit 2, as heretofore described.

It will be seen that the use of the air filters and 'air connections I85 permit control of air conditions, such as humidity, which may be of particular value in the event the specimens are plants. Likewise, temperature conditions may be controlled, either through the medium of the water jackets described, with their lines 50, I09, 31, I86, I81 or by heating one or more of the units by external means.

It will be further understood that, while the units may be sterilized by filling with an antiseptic solution and then draining said solution at I84 without admitting air other than by the air-filters, this method is limited by practical considerations to but a few applications because of the advantages of the sterilization by steam. One useof antiseptic sterilization is where it is desired to introduce some particular living material into the system for test of its effect on the specimen uninfiuenced by other or foreign matter. Thus it may be desired to introduce a particular kind of micro-organism into the system, in which event the same is sealed in a container, introduced into the desired unit for sterilization, the unit then be sterilized by introduction of the antiseptic solution, and the container of the micro-organisms opened only after the sterilizing operation. It will be obvious that most micro-organisms would be destroyed or killed by the high heat of steam, whereas the use of an antiseptic solution in the above manner will not so destroy the same.

I claim:

1. Apparatus of the character described, comprising a plurality of independent chambers, means interconnecting said chambers in sealed relation to form a sealed assembly, removable closure means interposed between said chambers and controlled by an operator from the exterior of said assembly, and means for sterilizing selected chambers while said assembly is sealed, said closure means being releasable to establish communication between said chambers without exposing the interior of any chamber to atmosphere.

2. Apparatus of the character described, comprising a pair of independent hermetically sealed chambers, a chambered member connecting said chambers, a closure at each end of said member for sealing said member from said chambers. means for selectively supplying steam to said member and chambers and sealed members in each chamber permitting manual access to said closures without destroying the sealed condition of the apparatus.

3. Apparatus of the character described, comprising a plurality of independent chambers,

means for connecting said chambers to provide an air-tight joint therebetween, a removable closure between said chambers, and flexible means carried by one chamber for aflordlng access to the interior of said chamber and to said connecting means and closure whereby said closure may be operated interiorly of said chamber, said means having an air-tight seal wit said chamber.

4. Apparatus of the character described comprising a sealed chamber having an opening therein, means for affording access to said chamber comprising an imperforate flexible member mounted in sealed relation at the periphery of said opening, said member being formed of material impenetrable by air and a removable closure for said opening disposed interiorly with respect to said access means.

5. Apparatus of the character described, comprising a container, having a pair of openings therein, a sealing closure for one of said openings, closure locking means within said container, a glove of material impervious to air mounted in the other opening in sealing relation at its perimeter, said glove being of a length suflicient to reach to said closure locking means.

6. Apparatus of the character described, comprising a pair of storage chambers, and a transfer chamber having an opening, elongated flexible means sealed at said opening and affording access to the interior of said chambers, means for interconnecting said chambers in a sealed assembly, and sealing closures for the inner ends of said storage chambers and accessible through said elongated flexible means for release to open said chambers into communication.

7. Apparatus of the character described comprising a plurality of chambers, means for interconnecting said chambers in sealed relation. means for opening said chambers into communication, at least one of said chambers having an opening therein, and a glove secured at its open end to the periphery of said opening, said glove being formed of material impenetrable by air and being of a length sufllcient to extend into connected chambers.

8. The combination with a chamber having an opening therein, of a glove secured at its open end to the periphery of said opening, a closure for said opening disposed inwardly of said glove, and means for releasably locking said closure.

9. In combination a chamber having an opening therein, a closure for said opening, means for locking said closure, a glove, and means for clamping the edge of said glove at the perimeter of said opening in sealed relation exteriorly of said closure.

10. In combination a chamber having an opening, an open ended member mounted in said opening in sealed relation, a closure at the inner end of said member, and a glove secured at its open end to the periphery of member in outwardly spaced relation to said closure.

11. In combinatioma chamber having an opening, an open ended member mounted on said chamber at said opening, a cllisure for the inner end of said member, means:avithin said member for operating said closure, a glove formed of air-impervious material sealed at its open end to the periphery of said member outwardly of said closure operating means, and a closure at the outer end of said member.

12. In combination, a chamber having an opening therein, an adapter ring secured to said chamber at the margin of said opening, an open ended membernhaving a sealed perimetral Joint with said adapter, a closure for the inner end of said member, and a glove secured at its open end to the inner periphery of said member.

13. In combination, a chamber having an opening therein, an open ended member having a sealed perimetral connection with said chamber at said opening, a releasable closure at the inner end of said member, a glove having a sealed perimetral connection with said member in outward- 1y spaced relation to said closure, and means for supplying steam to said member intermediate said closure and glove.

14. In combination, a chamber adapted to house a biological specimen and having an opening therein, an open ended water jacketed memher having a sealed perimetral connection with said chamber at said opening, a releasable closure at the inner end of said member, a flexible glove sealing the outer end of said member, and means for supplying steam to said member intermediate said glove and closure, said water jacket minimizing heat transfer to said chamber 15. In combination, a chamber adapted to house living matter and having an opening therei locking means for each closure, and a perimetral flange projecting exteriorly from said housing adjacent one end and spaced from the closure therefor adapted for connection with an open ended support through whose open said housing projects.

1'7. In biological apparatus, the combination with a chamber having an opening therein, a housing smaller than said chamber projecting through said opening, releasable means forming a sealed connection between said chamber and an intermediate portion of said housing, a closure for each end 01' said housing, and means within said chamber for operating the inner closure.

18. In biological apparatus, the combination with a chamber having spaced openings, of a glove formed of material impervious to air, means for mounting said glove to seal one of said openings, an open ended housing projecting through the other chamber opening, means forming a sealed joint between said chamber and housing, and a releasable closure at each end of said housing, said glove being of a length to reach said inner closure.

19. The construction defined in claim 18, and means for supplying steam to said housing, said inner closure being releasable to permit steam sterilizing of said chamber.

20. In biological apparatus, a pair of communicating chambers, one 0! said chambers having an opening. a rubber glove sealing said opening and afiording access to the interior of said chamber, areleasable closure for the inner end of the other chamber within reach of said glove, an exteriorly accessible releasable closure for said last named chamber, and means in said last named chamber tor sterilizing any material desired to be inserted in said first named chamber.

JAMES, A. REYNIERS.

Patent No. 2,244,092 Granted June 3, 1M1

JAMES A. REYNIERS The above entitled patent was extended January 8, 1952, under the provisions of the act of J une 30, 1950, for 2 years and 328 days from the expiration of the original term thereof.

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