WO2000000276A1

WO2000000276A1 - Method and device for treatment of whole blood

Info

Publication number: WO2000000276A1
Application number: PCT/SE1999/001184
Authority: WO
Inventors: Christian Freiburghaus; Britt-Marie Kjellberg
Original assignee: Fresenius Hemocare Beteiligungsges.Mbh
Priority date: 1998-06-30
Filing date: 1999-06-30
Publication date: 2000-01-06
Also published as: AU5074399A; SE9802322L; SE9802322D0; SE522921C2

Abstract

A method and a device for rinsing a product (6) which has been in contact with whole blood, such as an adsorption column for treatment of whole blood. The product is first rinsed with an isotonic solution (15) to remove the major part of the blood residue and thereafter with a hypotonic solution (19) to lyse remaining cells, which possibly may be performed with an isotonic solution. For an adsorption column (6), whole blood is passed through the column for adsorption of antibodies, such as immunoglobulin, on adsorption ligands immobilised in the column. The column is regenerated by passing an eluent solution (22) through the column. The elution step is preceded by rinsing the column with a hypotonic rinse solution (19), such as distilled water, so that possible remaining cells are lysed and rinsed away.

Description

TITLE

Method and device for treatment of whole blood

TECHNICAL FIELD

The present invention relates to a method and a device for rinsing of a product which has been in contact with blood. The invention particularly relates to a method and a device for regeneration of an adsorption column, which is used for whole blood.

BACKGROUND OF THE INVENTION

Dialysis, immunoadsorption and plasma exchange are extra- corporal treatment methods used in health care with the aim to eliminate unwanted substances in the blood of a patient. For dialysis, these unwanted substances are such that are normally excreted by the kidneys, which for these patients do not operate or operate unsatisfactorily.

Further examples of treatment methods, which include extracorporeal circuits are heart surgery, where pumps and oxygenators replace the heart and lung functions.

For immunoadsorption and plasma exchange, these substances are in most cases antibodies (immunoglobulin) .

Antibodies may be produced as an answer to an external agent (antigen) such as, for example, antibodies against coagulation factors at hemophilia, or antibodies against other individuals "transplantation antigens" (HLA-antibodies) or so called xeno antibodies. The above mentioned antibodies are not directly harmful to the body, but complicate or limit the possibilities for the individual to get adequate medical treatment .

In many other cases, the knowledge of the underlying agents which have caused the production of antibodies, is limited. It is through the effect of the antibody that the illness is recognised. Many of these are autoimmune conditions, such as, for example, Goodpasture ' s syndrome, Myasthenia Gravis, Guillain-Barre ' s syndrome, SLE and Wegener ' s Granulomatosis .

In certain cases, the antibody and antigen are known and have been characterised thoroughly, but in other cases only the method of treatment will reveal the fact that an antibody is causing the symptoms. In the latter case, it is probable that the antibodies participate in the mediation of the illness.

The methods for elimination of antibodies available today are, on the one hand, plasma exchange, where plasma unselectively is eliminated from the body and replaced with either plasma from healthy donors or albumin solution, and on the other hand, the more specific method of immunoadsorption. During immunoadsorption, immunoglobulin, or a specific substance, is eliminated by bonding to an affinity column having a suitable ligand, such as protein A or ligands aimed at a specific substance. It is possible to increase the capacity of the immunoadsorption system by utilising two columns which alternately are adsorbing and regenerating. This is preferably used for adsorption systems in a plasma environment, where plasma is passed through the column. The adsorption system used in whole blood environments are normally not regenerated. It is most often very complicated to get rid of all blood cells. Moreover, remaining blood cells may later be damaged during the regeneration (elution) often in a way that is harmful to the adsorbent. In US 4,708,714, a method and a device is disclosed for the treatment of blood or plasma by passing it through a column containing immobilised adsorption ligands. The adsorption ligands may be protein A, which is immobilised on a Sepharose gel, which is an agaros gel manufactured by Amersham Pharmacia Biotech AB . Such adsorption columns are sold by Excorim AB under the trademark "Immunosorba" . Through the method according to this patent publication, other substances are removed from the blood plasma through adsorption in the column.

When the adsorption medium is saturated, or at another point in time, an elution of the column is performed by letting an elution solution, having a low pH pass through the column. Meanwhile, a continued adsorption is performed in a second, parallel arranged column. Thereafter, the method is repeated.

Parts of the elution solution are collected for subsequent use of the separated antibodies.

In US 5,466,607 is disclosed a method and a device in which the above treatment and elution has been further improved .

As is mentioned above, these known methods are, in practice, utilised only on plasma, which first is separated from whole blood in a plasma filter and is passed through the column for adsorption of antibodies and thereafter reintroduced to the patient. If these methods were to be carried out on whole blood, it is very complicated to get rid of all blood cells and the remaining blood cells will be damaged afterwards during the regeneration (elution) and often in a way which is harmful to the adsorbent .

On the other hand, it is known to reuse dialysers by exposing them for a sterilisation method involving passing a rinsing solution through the dialyser and then introducing a sterilising agent, such as formaline. Another method of cleaning and sterilising a dialyser is described in EP 864 334, in which electrolysed alkaline water is introduced in the dialyser to clean it from proteins and other residual blood products, whereupon a rinse solution rinse the dialyser and electrolysed acidic water is introduced in the dialyser to achieve sterility between the treatments. However, a dialyser differs from a affinity column in that the blood contacting surface of a dialyser normally has a low affinity to blood products, like erythrocytes, while an affinity column has a high adherence . SUMMARY OF THE INVENTION

The object of the present invention is to propose a method and a device which makes it possible to rinse a product that has been in contact with whole blood.

The invention especially concerns a method and a device, which makes possible the repeated adsorption in an affinity column and subsequent regeneration, applied to whole blood. For this, a regeneration method is required which is adapted to usage with whole blood. The elution is usually preceded by a rinsing step in which plasma or whole blood in the column is displaced and reintroduced to the patient. Thereafter, the non-useable remains are rinsed away. Finally, the elution itself takes place using an elution solution having a lower pH, at which the bonding between the adsorption ligand and the antibody is removed.

As described above, during the use of whole blood, an effective elution of the column is prevented by possible remaining blood cells, which have not passed out of the column during the rinsing step. Remaining cells may be denatured during the elution phase and may deteriorate the adsorbent.

Therefore, it is suggested, according to the invention, to use a separate rinsing step to remove remaining cells. This rinsing step is performed using distilled water or another hypotonic solution. By using distilled water, the cells will be subjected to a strong osmotic gradient causing the cells to take up water and burst, so called lysing. It has unexpectedly been found that it is much easier to remove the remaining cell residues by rinsing, after the performance of such a lysing step.

We have found that such an interjacent cleaning step with distilled water which lyses the cells, provides a cleaner adsorbent which lasts much longer and that the regeneration time may be substantially shortened. This makes it possible, in clinical applications, to work with whole blood adsorbents which undergo a regeneration between treatments or even during treatments. The benefits are most pronounced in cases where a specific known antibody shall be eliminated or where a specific antigen or group of antigens are used during the elimination which means that only a limited part of the immunoglobulines will be removed. A good example is the elimination of xeno antibodies through the use of a mixture of specific carbohydrate ligands.

BRIEF DESCRIPTION OF THE DRAWINGS

Fig. 1 is a schematic diagram of an adsorption column set- up for antibody adsorption using whole blood.

Fig. 2A is a schematic diagram similar to Fig. 1 and shows a first step of the method of the invention.

Fig. 2B is a schematic diagram similar to Fig. 2A and shows a second step of the method of the invention. Fig. 3A is a schematic diagram similar to Fig. 2A and shows a third step of the method of the invention.

Fig. 3B is a schematic diagram similar to Fig. 3A and shows a fourth step of the method of the invention.

Fig. 3C is a schematic diagram similar to Fig. 3A and shows a fifth step of the method of the invention.

Fig. 3D is a schematic diagram similar to Fig. 3A and shows a sixth step of the method of the invention.

Fig. 4 is a schematic view and shows an apparatus in which the present invention may be used.

DETAILED DESCRIPTION OF EMBODIMENTS OF THE INVENTION

In Fig. 1 is shown a schematic view of a patient 1 (only an arm of the patient is shown) , who receives a whole blood treatment using a device according to the present invention. During the treatment, blood is removed from the patient via an artery needle or catheter to an artery tube 4, which passes an artery clamp 3. The artery tube 4 passes on through a first peristaltic blood pump 5 to a column 6, which contains a suitable adsorption ligand immobilised in a suitable way in the column for adsorption of antibodies and/or other substances.

The purified blood passes from the column 6 in a vein tube 7 through a drip chamber or air guard 8 having a pressure monitoring device 9, and via a vein clamp 10 back to the patient 1 via a vein needle or catheter 11. The whole set of tubes is sterilised before use.

Citrate may be added for anticoagulation from a container 12 via a tube 13 which passes a second peristaltic citrate pump 14 and connects to the artery tube 4 at or just after the artery needle 2.

The treatment continues during a predetermined time, for example until the column is saturated and no further adsorption takes place. Thereafter, the device is connected into a displacing position as shown in Fig. 2A.

A container 15 for a rinsing solution, such as sterile physiological sodium chloride solution, is connected via a tube 16 to the artery tube 4, which has been disconnected from the arterial needle 2. Rinsing solution is pumped in a first step via the artery clamp 3 and the blood pump 5 through the column 6 to displace the blood back to the patient. When all the blood has been displaced, the device is switched to a regeneration position.

The device is further reconnected so that the vein tube 7 is connected to a discharge or drain 17a, in a second step as shown in Fig. 2B. Further rinsing solution is passed through the tube 16 and column 6 to drain 17a.

Thence, a container 17 with sterile neutral buffer solution is connected using a tube 18 via the vein clamp 10 and the blood pump 5.

A container 19 with distilled water is connected using a tube 20 via the vein clamp 10 to the citrate pump 14 and onwards via a tube 21 to the column 6.

A container 22 with eluent is connected using a tube 23 via the artery clamp 3 and the citrate pump 14.

The device operates in the following manner in the regeneration position, as shown in Figs. 3A, 3B, 3C and 3D.

In the third step, the column 6 is rinsed, using the sodium chloride solution from the container 15 via the tube 16, the artery clamp 3 and the blood pump 5, in the same way as during the second step, and the drain of the column is lead to the drain 17a, as shown in Fig. 3A. In a fourth step, distilled water is passed from container 19 via the tube 20, the vein clamp 10, the citrate pump 14 to the column 6 to lyse possible remaining cells and rinse the residue to the drain 17a, as shown in Fig. 3B In a fifth step, eluent is passed from container 22 via the tube 23, the artery clamp 3, the citrate pump 14 to the column 6 to elute the adsorption ligands and to free adsorbed antibodies or other substances, as shown in Fig. 3C. The elution is more efficient since the column is completely cleaned in the previous steps.

In a sixth step, a neutral buffer solution is passed from the container 17 via the tube 18, the vein clamp 10, the blood pump 5 to the column 6, as shown in Fig. 3D, in order to neutralise the column and to put it in order for treatment in a new treatment position according to Fig. 1.

As is apparent from what has been described above, switching between the four regeneration steps may be performed completely automatically by actuation of the artery clamp 3, the vein clamp 10, the blood pump 5 and the citrate pump 14 in a predetermined sequence .

Switching between treatment position, flushing position and regeneration position may be performed using valves or by rearrangement of tubes according to a predetermined pattern. Switching between the different positions is suitably performed manually by observing whether blood or salt solution is present in the tubes .

Another embodiment of the present invention used on the device according to US 5,466,607 is shown in Fig. 4.

In Fig. 4, P indicates a connection to a patient. Normally plasma is obtained from the patient, but according to the present invention, it is whole blood. A pump PP pumps the blood to a valve V2 and further to a drip chamber ADB and an adsorption column B. From there, the blood passes via a sensor pDB and a valve V4 and a pH-sensor pHP and a valve V6 back to the patient P' .

A second adsorption column A is connected in a regeneration circuit, comprising a container for an acidic eluent solution AC, which is supplied to the column A via a pump PA and a valve VI and a drip chamber ADA. From there, the eluent solution passes via a plasma sensor pDA, a valve V3 , a sensor UV and a valve V5 to a container F for collecting the eluent fraction. Further, there is a container for buffer solution B, such as physiological sodium chloride solution (common salt) , which is used to displace the blood in the column before elution and possibly to displace the acidic eluent solution before addition of new blood for adsorption. This process is regulated by the valves VI and V2 , together with the pump PB.

A container W for non-usable waste solution, which normally will be discarded, may be connected instead of the container F for the fraction or the connection P' for the patient, via the valves V5 and V6. The valves V3 and V4 are actuated according to the valves VI and V2 , but with a delay, which is determined by the sensors pDA and pDB and regulated by a microprocessor MC .

For the exact operation of this apparatus, please refer to the patent publication US 5,466,607, which is hereby incorporated by reference in the present application.

As is evident from Fig. 4, a further container H is used according to the present invention, containing a hypotonic solution, such as distilled water. The solution is pumped from the container H using a pump PH and reaches the valve VI and is connected in parallel with the acidic eluent solution or the buffer solution.

The regeneration step is performed in the following way according to the present invention. When the valves VI and V2 are switched to the position showed in Fig. 4, the column contains whole blood, which has been obtained from the patient connection P.

Firstly, the buffer pump BP is actuated and will pump buffer solution through the column A to thereby displace whole blood from the column A. The blood is returned to the patient via P', until the sensor pDA deactivates the valve V3.

Thereafter, the solution will pass to the container W because the valve V5 is actuated. The column A is flushed to remove cells and other products from the column. There is, although, always red and white blood cells left, which have got stuck between the Sepharose balls or which stick to cracks or other small irregularities in the column. These cells prevent the following elution and cause the column to not be fully regenerated.

After this, the pump PB is stopped and the pump PH is actuated to pump hypotonic solution through the column A. The hypotonic solution causes the cells to be subjected to a strong osmotic gradient, so that the cells will burst and be lysed. Thereafter, it is much easier to flush out the cell residue, which is collected in the container W.

Thereafter, the column is eluted in a known way by actuating the pump PA and the usable fraction is collected in the container F. Finally, the pump PB is actuated to displace the eluent solution from the column A (to the waste container W) before whole blood again is connected to this column.

The above examples show that the invention may be used in a partly manual device or in a fully automatic device for whole blood adsorption. Further usage of the invention for other prior known devices should be apparent to a person skilled in the art, considering the above description.

The invention has been described above with reference to preferred embodiments of the invention. A person skilled in the art will realise that further combinations are possible. Such modifications, which are apparent to the person skilled in the art are intended to be encompassed within the scope of the invention. The invention is solely limited by the appended patent claims .

Claims

PATENT CLAIMS

1. A method for rinsing of a product which has been^" in contact with blood, characterised in that the product is rinsed with a hypotonic solution so that possible remaining cells are lysed and rinsed away.

2. The method according to claim 1, characterised in that the product is first rinsed with an isotonic solution to remove as many cells as possible, then with a hypotonic solution to lyse and rinse away the remaining cells, and finally once more with an isotonic solution for final rinsing.

3. A method for treatment of whole blood, comprising passing whole blood through an adsorption column to adsorb substances in the blood liquid, such as antibodies, on immobilised adsorption ligands in the column, interruption of the passage of whole blood through the column to regenerate the column by elution of the adsorbed substances by means of passing an eluent solution through the column, repeating the previous step until the treatment is finished, characterised in that the elution is preceded by a rinsing of the column with a hypotonic solution so that possible remaining cells are lysed and rinsed away.

4. The method according to claim 3, characterised in that the regeneration is performed with the following steps: rinsing of the column with an isotonic solution to displace the whole blood for restoration to the patient, continued rinsing of the column with an isotonic solution to flush the column clean from cells and other remaining lose products, whereupon the rinsing solution is discarded, rinsing of the column with a hypotonic solution for continued cleaning of the column whilst simultaneously lysing possible remaining cells, elution of the column with an eluent solution, for example having a low pH, to free the adsorbed substances, which may be collected in a container, rinsing of the column with a buffer solution to put it in order for repeated adsorption.

5. The method according to claims 3 or 4 , characterised in that the regeneration step is performed simultaneously with continued adsorption in a step arranged in parallel in a multicolumn system.

6. A device for carrying out the method according to claim 1, for rinsing a product which has been in contact with blood, characterised by a source (19) of a hypotonic solution and a device (10, 20, 14) for rinsing the product (6) with the hypotonic solution to lyse and rinse away possible remaining cells from the product.

7. The device according to claim 6, characterised by a source (15) of isotonic solution and a device (16, 5) for rinsing the product with the isotonic solution, before and/or after the rinse with the hypotonic solution, to rinse away remaining cells.

8. A device for carrying out the method according to claim 3, for the treatment of whole blood, comprising an adsorption column (6) for adsorption of substances such as antibodies, for example immunoglobulin, on adsorption ligands immobilised in the column, during the passage of whole blood therethrough, a regeneration device (22, 23, 14) for the regeneration of the column by elution of the adsorbed substances by passing an eluent solution through the column, characterised in that the regeneration device comprises a device (19, 20, 14) for rinsing of the column with a hypotonic rinse solution so that possible remaining cells are lysed and rinsed away.

9. The device according to claim 8, characterised in that the regeneration device comprises devices for: rinsing (15, 16, 5) of the column with an isotonic solution to displace whole blood for restoration to the patient , continued rinsing (15, 16, 5) of the column with an isotonic solution to flush the column clean from cells and other remaining lose pollutants, whereafter the rinsing solution is discarded, rinsing (19, 20, 14) of the column with a hypotonic solution for continued cleaning of the column whilst simultaneously lysing possible remaining cells, elution (22, 23, 14) of the column with an eluent solution, for example having a low pH, to free the adsorbed substances, which may be collected in a container, rinsing (17, 18, 5) of the column with a buffer solution to put it in order for repeated adsorption.

10. The device according to claims 8 or 9 , characterised by a multicolumn system (A, B) so that the regeneration step is performed simultaneously with continued adsorption in a step arranged in parallel.