IL34815A - Electrophoresis device - Google Patents

Description

Electrophoresis Device BAR-I'LAK OTTOiSXTY Inventor: Dr. Ram Avtalion The present invention concerns an electrophoresis device.

Electrophoresis is nowadays a standard laboratory technique applied in particular for chemical, physical and weight biological studies of high-molecular rganic substances such as proteins and polypeptides, and is used for both analytical and preparative purposes.

In accordance with one division, electrophoresis methods and devices are divided into vertical and horizontal. In the former, the suspended material migrates along a vertical path and the latter along a horizontal path. By a further division, the existing electrophoresis methods and devices are divided into analytical and preparative. Hitherto a special device has been needed for each purpose and technique. Thus there exist preparative vertical electrophoresis devices, analytical vertical electrophoresis devices and analytical horizontal electrophoresis devices.

Each electrophoresis device is associated with a considerable amount of electronic hardware serving for detection of different fractions, control of fraction collectors and the like. In consequence, each electrophoresis device forms part of an assembly that is both bulky and expensive. Each of these properties constitutes a considerable drawback when having regard to the fact that any investigator in the field will have to be equipped at least with one preparative and one analytical electrophoresis device, and preferably with two analjfclcal devices , one for vertical and the other for horizontal electrophoresis since there are cases where the on© and other cases where the other type is to be preferred.

- Hereinafter the said gel-electrophoresis cell will be referred to for short as "cell".

When the device according to the invention is used for analytical horizontal electrophoresis, the cell will be inoperative and the media for the horizontal electrophoresis are placed on top of the horizontal bridge member.

In case of vertical analytical electrophoresis with the aid of the cell, the said horizontal elution channel will not be formed and the cell will be filled with two superimposed gel bodies fully bearing on each other, the upper one serving as medium for electrophosetlc migration and the lower one fulfilling the double function of serving as a carrier base for the upper one and as means for establishing an electrolytic connection between the upper gel body and the electrolyte below and around the cell. This connection is made possible by the liquid permeability of the lower gel body and the opening(s) in the bottom of the cell.

Where the device is to serve for preparative electrophoresis, a passage is formed between said superimposed gel bodies. Moreover, in that case the upper gel body will preferably rest at least partly on brackets of the cell, especially introduced for the purpose, and the main function of the lower gel body remains for establishing the electrolytic : connection as specified.

During operation of the device tot preparative electrophoresis, said passage serves as elution chamber and any material arriving at the passage is eluted by the elution liquid flowing therethrough. V/ith the aid of conventionaljelectronic detector means associated with fraction: collector means, it is possible Fig. 12 is a side view of the cell according to Pigs. 6 to 9 in the semi-assembled state for the formation of an elution chamber by gel casting; Fig, 13 is another side view of the wall according to Fig. 10; Fig. 14 is a fragmentary elevation of a feeder and an accessory funnel device; Fig. 15 is a side view of another embodiment of a feeder; Fig. 16 is a fragmentary elevation of the feeder according to Fig. 15 and an accessory funnel device; Fig. 17 is a fragmentary perspective view of the assembled cell ready for casting, with said means for causing the formation of an elution chamber by casting in position; Fig. 18 is a fragmentary elevation of the cell corresponding to Fig. 17; Fig. 19 is an elevation of means for insertion in an assembled cell to cause the formation therein of an elution chamber by gel casting; Fig. 20 is a section along line XX-XX of Fig. 19 drawn to a larger scale ; Fig. 21 is a cross-section through the device according to Figs. 1 and 3 in preparative operation; and Fig. 22 is a cross-section through the cell of a device according to Figs. 1 and 3 in analytical operation.

The casing of the device illustrated in Fig, 1 comprisee a rectangular prismatic box 1 and a removable cover 2. A number of recesses 3 at the upper edges of the lateral walls 4 of box 1 serve for leading into. the box electric wires and flexible tubes By means of partitions 6 and 7 co-extensive with the longitudinal side walls 5» box 1 is subdivided into a main, operational compartment 8 of inverted L-like cross-sectional shape, and a storage compartment 9 serving for storing therein the various accessories of the device and accessible by means of a sliding closure 10. The horizontal partition 6 serves as a support for a trough 12 which serves as the first electrode compartment (see also Figs. 3> 4 and 21 ) and houses one of the electrodes, which is in the form of a stretched platinum wire 19 mounted on an insert 18. Wire 9 is connected to a flexible wire (not shown) fo connection to one terminal of a DC source.

The remaining space of the operational compartment 8 serves as second electrode the second electrode compartment 13.whieh/is in the form of a stretched platinum wire 15 mounted on an Insert 14 and connected to a flexible wire (not shown) for connection to the other terminal of a DO source.

The Inner components of the device according to Pig. 1 are shown in an assembled state in Pigs. 3 and 4· These parts comprise the said trough 12 with said insert 18 (see also Pig. 21 an electrophoresis cell 17 adapted for insertion into the second electrode chamber 13 and said insert 14 (see also Figs. 1 and 21). When cell 17 is in position it forms part of chamber 13 in which it is located.

Amon the inner components of the device there appears a horizontal bridge member 20 fitted with spacer ribs 21 and having longitudinal slots 22. Ribs 21 serve for the correct positioning and fixation of the bridge member. Bridge member 20 the upper edges of rests on/trough 1 and cell 17 as shown in Pigs. 3 and 21.

As seen from Fi s. and 2 one of the lon itudinal - - When carrying out vertical electrophoresis, "both analytical and preparative, the electrolytic connectIon--between trough 12 and cell 17 is established by a pair :of sheets 23 made of pliable , liquid, absorbin material ' thfitt is inert to the electrolyte employed1 for electrophoresis and capable of transferring electro-lyte from one compartment to the other by a capillary sucking effect. Example ;are filter paper, cellular synthetic materials and the like. One of sheets 23 dips intp-. rough 12 through the cpprdinated slot 22 and the other one dips ; into cell 17 through the slot 22 associated therewith. As is urther seen, the two sheets 23 overlap and: i this wa the electrplytic connection between the two electrode compartments is established. Alternatively, it is also possible to establish direct connection between the two- electrode compartments :by providing s table holes, b res Or quite generally liquid permeable passages through the partition between the two compartments..

When the device is used or horizontal electrophoresis the cell 17 is not required but need not be removed. In this case carriers 2 for the electrophoresis media will be placed pm bridge member .20, as shown, in Fig. 2, arid?, two tongues 250 of pliable, liquid absorbing material that is inert to the electrolyte employed^ for electrophoresis and capable of transferring electrolyte from one compartment to. the other, are associated with each carrier 24 and depend from each carrier, into the associated electrode chamber., ; Cell 17 is. fitted at its lower part with flexible tubes and 26 fpr the introduction and withdrawa pf e utio liquid, as will be e plained further below. · : The structure of cell 17 is more closely shown in Figs.

Cell 17 Is in itself a dismountable assembly of interlocking members comprising side walls 30 held together through Interlocking engagement by means of a first connector rib 31 and a second connector rib 32. Each of side walls 30 comprises in its upper portion a longitudinal recess so that the walls receive between them in fixed position a feeder 33, as shown more particularly i Figs. 8 and 9· The feeder 33 according to Fig. 5 comprises a plurality of cylindrical bores 34 while the feeder 33£, according to Fig. 6, comprises a plurality of oblong slots 34a. The feeders are exchangeable and cell 17 may be itted with adequate feeders according to requirements. When the cell is filled with gel bodies, as will be explained belo „ the width of the bodies determined by the distance between the side walls o the cell correspond to the width of the openings 34 or 34a, se the case may be. This in tur is determined by design Of connector ribs 31 » 32. Thus in Fig. 5 the width of the central parts 131 and 132 of members 31, 32 corresponds to the diameter of bores 34» while the central parts 131 and 132a, in Fig. 6 have the same width as slots 34..

The embodiment of Fig. 6 is suitable for cases of analytical electrophoresis in which the medium is after the run sliced into thirty long strips.

At one lateral side the lower portions Of the side wall 30 are so shaped to form together with the connector rib 31 a circular socket/ (see Fig. Θ) adapted to receive a plug 3.6 for connection of the flexible hose ¾6. In a similar way side walls elution chamber is needed when the device is used for repa a^ve electrophoresis, as will be explained further below«.

For still better understanding, the design of each side wall 30 and of the connector ribs 31 and 32 is shown in Pigs, 10, 11 and 1 . Fig. 10 shows the inner face of a side wall 30 and depicts the following: A profiled groove 41 for cooperation with connector rib 31, a profiled groove 42 for cooperation with connector rib 32, a recess 43 for the accommodation of the feeder 33 and a profiled groove 44 for oooperation with a ledge 38 either of the first or second kind.. In Fig. 10 such a ledge is shown in the oourse of being positioned by sliding from left to right. The ledge shown in Fig. 10 is of the second kind and only part of it is shown, the remainder, on the left-hand side,being broken away. Fig. 10 further depicts a recess 45 which together with an adequately shaped recess in the connector rib and an identical recess in the other side wall 30 forms in the assembled oell the socket 35 (see Fig. 8) and on the opposite side a recess 47 which together with a recess in connector rib 32 and an identical recess in the other side wall 30 forms in the assembled cell the circular sooket 37 (see Fig. 9) .

I is further seen from Fig. 10 that each side wall 30 comprises in its lower region a longitudinal groove 50 which serve for anchorage of the lower gel body.

Fig. 11 is a side elevation of the connector rib 31 whose plan view is given in Figs. 5 and 6 and front elevation in Fig. 8. In its lower part rib 31 comprises a recess 48 which in the assembled cell together with the recesses 45 of the flanking side walls 30 forms the cylindrical sooket 35· Fig. 11 sl t Pig. 13 ia a side elevation of the connector rib 32 whose plane view and front elevation are given in Figs. 5 and 6 and Pig. 8 respectively. As seen, the rib comprises a recess 49 which together with the recess 47 of the flanking side walls 30 forms the cylindrical socket 37· Fi . 1 3 further depicts the bore 40 which is stepped, as shown, for the accommodation of a correspondingly shaped plug 138.

For the assembl of the cell each side wall 30 is first fitted with its ledge 38 by sliding it into the profiled groove 44. Thereafter the feeder 33 is positioned into the recess 43 of either of the two side walls and the second side wall 3 is then positioned so that the feeder is sandwiched between them* The resulting sandwich is ready for receiving connector ribs 31 and 32 which are slid in from above and arrested in their final positions by means of adequate stops. The resulting assembly 1B ready for gel casting.

The joints between connector ribs 31 and 32 on the one hand and side walls 30 on the other hand are designed to be liquid-tight in order to prevent the 1ε kage of electrolyte through them. This tightness Is achieved in the embodiments hereinbefore described by the profilation of the ribs and the grooves by which they are received. A similar effect may be achieved by gasketlng, in which case th® ribs and grooves may be of a simple design which may be advantageous from a manufacturing point of view.

After completion of an electrophoresis operation the cell is dismantled into its components and the gel bodies are removed therefrom. Where the electrophoresis was analytical, the gel may be sliced and submitted to further examination as known S In Pigs. 7, 8 and 9 the assembled cell is shown with ledges 38 of the first kind. These ledges are, as mentioned, so dimensioned that when positioned in groove 44 each such le¾e is flush with the inner face of wall 30* In this way a continuous inner space is formed within the cell for the casting of two continuous bodies of gel fully bearing on each other. In this case there is noti formed an elution chamber and the cell is adapted for application for analytical electrophoresis* The upper gel body that serves as medium for the electro-phoresisand the lower gel body serving as support are both formed in situ by casting gellifylng liquids. The lower auppr-rting gel body is best cast by dipping the cell in a bath (see Fig. 17) while the upper gel body is cast by pouring gellifying liquid through feeder 33» Obviously the liquid that forms the upper gel body can only be cast after the liquid that forms the lower gel body has fully occupied its space.

For pouring the gellifying liquid through feeder 33 there is provided a funnel member for association therewith comprising a plurality of outlets corresponding in number and shape to opening 34 or 34a of feeder 33· As shown In Fig. 14, the feeder 3¾is of the kind show in Fig. 6, i.e. with a plurality of slots 34a,. The funnel device 51& comprises a manifold 52a. having a trough 5½ and a plurality Of depending hollow fingers 53a Each finger 53a, is adapted for a snug fit in a slo 34. and the interior bore of each finger then extends from manifold 52a, into slot 34.. For the casting operation the funnel 51 . is inserted into engagement with the feeder 33a BO that each slot 34,§ receives one finger 53. which penetrates to a level slightly above the lower edge Of bore 34&. as shown by dashed lines in the simultaneously at an equal rate through all the fingers 53a. and through the associated slots 34a into the inner space of the cell 17. When the gellified liquid reaches the level of the lower edge of opening 34a,» the casting operation is interrupted and the funnel member 51 , is extracted from the feeder 33a..

The funnel member of Pigs. 15 and 16, the former being a side view, is in principle of the samedesign, fingers 53 bein adapted for cooperation with the cylindrical bores 34 of the feeder 33 according to Pigs. 5 and 7.

Pig. 12 shows a cell 1 ready for casting to produce an elution chamber with the connector rib 31 being removed for clarity of illustration. It is seen from that figure that each ledge 38 of the second kind comprises a portion which projects beyond the inner face of the associated side wall 30 to form a longitudinal bracket. The two brackets ace each other leaving between them a narrow slit adapted to accommodate the blade of a plunger 55. Plunger 55 comprises a lower portion of nearly circular cross-sectional shape.

In practice when the connector rib 31 is in position the plunger 55 is inserted through slot 39 (see also Pig. 8). Cell 17 with the plunger 55 in position is shown in Pigs. 17 and 18 which are respectively a fragmentary perspective view and a fragmentary elevation of the cell. As is seen more closely from Pig. 19 and Fig. 20, the plunger 55 oomprises a supporting rod 55 and the right-hand side end of that rod is received by the innermost portion of bore 40 (see Pigs. 9 and 13) . It is further seen from Pigs. 19 and 20 that the plunger comprises a number of vents 56 which permit the venting of the air during the oaeting opera cell 17 rest on terminal bosses (not seen in the section). It is further seen that an elution chamber 60 is formed betvieen the facing brackets constituted by the jutting portions of the ledges 38 of the second kind (see also Figs. 12, 17» 18) separating the upper gel body 62 from the lower one 63. !he charges to be electrophorlzed are introduced through feeder 33 and the electrolytic connection between compartments 12 and 13 (the latter including gel 17) is established by sheets 23. which ensure an electrolyte transfer from compartment 1 to compartment 13* made possible by the fact that the liquid in the former is higher than in the latter indicated in each case by a dashed line. T e material arriving in chanber- 60 is eluted by the elution liquid flowing therethrough. The electrolytic connection between gel bodies 62 and 63 is established by the elution liquid flowing through elution chamber 60.

Pig. 22 is a fragmentary seotion through a devioe according to Pigs. 1 to 20, showing the cell 17 thereof prepared for analytical electrophoresis. It is seen that in this case ledges 38 are of the first kind and gel body - 62 fully bears on gel body 63· Gel body thus fulfils the double function of serving as support or gel body 62 and of providing direct electrolytic contact with the latter. how Prom the above description it becomes evident/the device according to the invention may serve without any mechanical changes and adaptations for any of the operations vertical preparative electrophoresis, vertical analytical electrophoresis and horizontal electrophoresis.

Claims (1)

1. insufficientOCRQuality