CA1199577A

CA1199577A - Liquid mixture for taking up and stabilising gas bubbles, and its manufacture and use as a contrast medium for ultrasonic diagnosis

Info

Publication number: CA1199577A
Application number: CA000412931A
Authority: CA
Inventors: Jurgen Hilmann; Rolf-Rudiger Hoffmann; Wolfgang Mutzel; Ingfried Zimmermann
Original assignee: Schering AG
Current assignee: Bayer Pharma AG
Priority date: 1981-10-16
Filing date: 1982-10-06
Publication date: 1986-01-21
Also published as: ZA827577B; DK160741C; JPS5879930A; AU558152B2; NO823452L; EP0077752B1; NO158328C; IE822490L; EP0077752A2; US4466442A; EP0077752B2; ATE18356T1; FI81008B; DE3141641A1; IE55051B1; FI823474L; DK160741B; FI81008C; AU8916382A; DK455782A

Abstract

A B S T R A C T

Liquid mixture for taking up and stabilising gas bubbles, and its manufacture and use as a contrast medium for ultrasonic diagnosis.

The invention is concerned with an ultrasonic contrast agent for use in diagnosis of the human or animal body, which comprises a solution in an aqueous liquid carrier of (a) a tenside, and (b) a viscosity-increasing substance the solution having dispersed in it a plurality of micro-bubbles containing a physiologically tolerable gas, especially sterile air, oxygen, carbon dioxide, nitrogen, helium and/or argon.
The invention also relates to a liquid medium for use in preparing the ultrasonic contrast agent, to a diagnostic kit and to a method of diagnosis of the human or animal body wherein the contrast agent is injected intravascularly.

Description

35~'7 The invention i~ concerned with ultrasonic 5 contra~t agent~ for use in the ultra~onic diagnosis of the human or animal body, and al~o with liquid media used in preparing such contra~t agent~ and which media are capable of taking up and stabilising micro-bubbles of air or other physiologically tolerable 10 ga~.
It is generally known that the image contrast in ultrasonic diagnosis iq increa~ed by the presence of ga~ bubble~ in the fluid, usually blood, pa~sing through the part of the body under examination. For 15 thi~ purpose it i~ possible to manufacture these gas bubbles outside the part under examination and to inject them into the bloodstream. l~i5 iS po~sible, for e~ample, by vigorou~ly shaking a liquid ~olution, such as common salt ~olution, a dye ~olution or previou~ly 20 removed blood, in order to produce the bubbles, then injecting lt and carrying out the ultrasonic examination.
For example, Feigenbaum et al., in the article ~Identification of Ultrasound Echoes From the Left Ventricle of the Heart Through the Vqe of Injection~
25 of Indo~yanine Green~ (Circulation, Volume XLI, April ~970) report~ on the production of echoe~ by means of ~AJ, f~

5~'7 -- 2 -- _ ga~ bubble~ in the left ventricle of the heart, as does also Gramiak et al., (Radiology 100, 415-4~8, ~971 ) .
Another proce3~ for producin~ micro-bubbles of dete~mined size is described in the report UNon-S Inva9ive Asse~ment of Pulmonary Hypertension U~ing theBubble Ultra~onic RP~onance Pre~sure (BURP) Method~
(Report No. HR-62917-1A, April ~977, Divi~ion of Lung ~isea~es, National ~eart, Lung and Blood Institute).
U.S. Patent No. 4 265 251 describe~ the u~e and 10 manufacture of micro-bubble~ with a saccharide covering that can be produced with reproducible size distribution by using an expensive complicated apparatus. The diq-advantages of this proce~s consist, on the one hand, in the fact that shortly before use the micro-bubbles 15 are mixed with the carrier in an open container, as a result of which sterility and freedom from pyrogens are not en~ured. On the other hand, the manufacture gives rise to high costs a~ a result of the complex technique3. A further U.S. Patent No. 4 276 885, for 20 example, describes a process for manufacturing ga~
micro-bubbles having a gelatine membrane and u~ing a gellable medium as carrier for the~e micro-bubbles, ~o that the gas bubble~ can be frozen by cooling and freed again by heating when required~
The disadvantage of this method is that a su~-pension produced in thi~ manner cannot be ~terilised, ~ince in the case of heat ~terilisation the micro-7'~

- 3 - _ bubbles are not stable and they are separated or de~troyed al~o when sterilised by filtration. In addition, the gelatine preparations always present the risk of anaphylaxi 3 ~
It i~ an ob~ect of the pr~sent invention to provide an ultra~onic contra~t agent in which the above-mentioned di3advantage~ are overcomeO
The present invention provides an ultrasonic csntrast agent for use in diagnosi~ of the human or animal body, w~ich comprise~ a 301ution in an aqueous liquid carrier of ta) a ten~ide, and (b) a visco~ity~increa~ing ~ub~tance, the solution having dispersed in it a plurality of ~5 micro-bubbles containing a physiologically tolerable ga~. Advantageously the solution contains from 0.01 -10 % by weight o the ten3ide (a) and from 0.5 ~ 50 %
by weight of the viscosity-increasing ~ubstance lb~.
The invention also provides a liquid medium for use in making up the ultra~onic contrast agent which comprise~
(i) a 301ution of one or more ten~ide~ in an aqueous liquid carrier, in admixture or , conjunction with 5 ~ii) a ~olution of one or more vi~co~ity-increa~ing substance~ in an aqueous liquid carrier.

5~'~
- 4 - _ The development of the micro-bubbles in the ~olution shortly ~efore use can be efected by widely varying methods, for example:
1) by drawing up by ~uction in an in~ection syringe or the like air or other p~ly~io-logically tolerable gas at the same time as drawing up the ~olution containing the tenside and visco~ity-increasing ~ubstance and repeating the drawing-up and squirting-~o out operations qeveral time~ ~Ro-called pu~pingt under ~terile conaitions;
23 by first of all separately manufacturing and then heat-sterilising a solution con-taining the tenside and a solution contain-~5 ing the vi~cosity-increa~ing ~ubstance, drawing up the first solution in a syringe or the like and then injecting this first solution into the second, the second ~olution being in a sterile vessel together with a physiologically tolerable ga~; and 3) by first of all separately manufacturing and then sterilising a solution of the tenqide that contains from 0.05 to 5 % by weight of a phy~iologically tolerable 2S carbonic acid salt, and a solution of the visco~ity-increasing substance that contain~ an amount, equivalent to the amount of carkonic acid salt, of phy~iologic-ally tolerable acid, and mixing them together under ~terile conditions ~hortly before use.
The mixing of the two solution~ ~an be effected by any method which achieves a vigorous swirling action, for example by Mechanical stirring, by ultra-sonic means, or by drawing-up one solution into an injection ~yringe and emptying the content~ of thi~
~0 syringe into the ~econd ~olution using the highest practicable pre~ure and a high discharge ~peed and then ~haking vigorously, it being necessary to en~ure sterile conditions for all the mixing operations.
For example, there i~ u~ed for the mixing operation a ve~sel that permits sterile condition~ and i~ of adequate size and, after recei~ing the second solution, still has a sufficiently large ga~ space for the ~ubsequent vigorous shaking. I~ is preferable to use for thi~ purpose sufficiently large multi-vials or ampoule~ having a clo~ure that can be pierced by an in~ection needle thus enabling in~ection of the solution, the subsequent mixing and the removal of the liquid medium containing micro-bu~ble~ dispersed in it, without being opened.
If nece~sary, in~tead of ~terile air the micro-bubble~ can be filled with other phy~iologically tolerable ~terile ga~e~ ~uch, for example, a~ c~rbon 1 ~ '7'^~

dioxide, o~ygen, nitroyen, noble ga~e~ or mixtures of two or more of 3uch ga~e~, ~terile air, carbon dioxide and/or oxyg~n being preferred. For thi~ purpo~e, the air i~ displaced from the solution of the tenaide or - 5 ten~ide~ and/or from the solution of the vi~cosity-increasing ~ub~tance or ~ubstance~, by treatment with the de~ired gaR and the two ~olution~ are mixed in one of the above-de~cribed manner~ in a multi-~ial that is filled with th~ de~ired ga~ or ga~ mixture.
If carbon dioxide i~ desired as the ga~, it i~
al~o po~ible to produce the ga~ during the mixing operation by adding to either the tenside ~olution or the ~olution of the viscosity-increasing Yubstance, from 1 to 3 % by weight of a physiologically tolerable acid ~uch, for example, as hydrochloric acid, tartaric acid, citric acid or an acidic salt of phosphoric acid, in the form of a dilute aqueous 301ution, and adding to the second ~olution the equivalent amount of a carbonic acid salt such, for example, as an alkali metal bicarbonate, for example, sodium bicarbonate or ammonium bicarbonate, in the form of a dilute aqueou~
301ution~
After the production of the micro-bubble~ by one of the above-mentioned procedure~ or by an equivalent procedure, the solution~ are prepared for intravenou~
or intra-arterial administration as an ultrasonic contra~t agent for ultrasonic diagno~

- 7 - ~ _ There i~ thus made available a contrast agent for ultrasonic diagno~i~ that i~ free of ~olid particles and i~ sterile. Furthermore, the in~ection of a ~mall volume of the liquid mixture according to ~he invention S ef fect~ an excellent increase in the ultrasonic contra~t.
In order to obtain the increaqe in ultrasonic contra~t obkain~d after intravenou~ injection of only 0.~ ml of the contrast agent according to the invention, which contain3, for exampl~, 1 % by weight of polyoxy-~O ethylenepolyoxypropylene polymer having a molecularweight of from 6800 to 8975 (Pluronic ~ F 68) and 4 % by weight of glucose in water, it i~ advantageou~
to u~e 3 ml of the suspension described in U.S. Patent No~ 4 276 885.
~oth non ionic and ionic tensides are ~uitable a~ tenside~O As non-ionic tensides there may b~
mentioned: lecithin3, lecithin fractions and their modification product~, polyoxyethylene fatty acid e~ter~, such a~ polyoxyethylene fatty alcohol ether~, polyoxyethylated sorbitan fatty acid esters, glycerine polyethylene glycol oxy~tearate, oxyethylated rhizinoleic glycerol ester, ethoxy-substituted soya sterols, ethoxy-3ub3tituted castor oil3 a~d their hydrogenated derivative3, chole~terol, and polyoxyethylenepolyoxy-propylene polymers, polyoxyethylene fatty acid stearate3and polyoxyethylenepolyoxypropylene polymers having a molecular weight of 6800 - 8g75, ~3300 and ~6250 ~eing - 8 - _ preferredO ~ ercentages relate to weight.
A~ ionic tensides ~here corne into consideration:
quaternary a~mmonium ba~e~, 30dium lauryl sulphate and ~odium dioctyl ~ulphosuccinate.
S q'he contra~t agent solution of the invention may contain from 0.01 to 10 %, preerably from 0.5 to 5 %, by weight o~ the tenside or mixture of ten~ides~
As ~iscosity-increasing substances there come into considerationO mono- or poly-saccharides such, for example, as glucose, laevulose, galactose, lacto~e, sorbitol, mannitol, xylitol, saccharose or dextrans, cyclodextrine3, hydroxyethyl starch and polyols. As polyols there may be used glycerine, polyglycols, inulin and 1,2-propanediol. There may also ~e used as the viscosity-increa~ing substance proteins, protein-like substances, amino acids or blood substitute substances such, for example, as plasma proteins, gelatine, oxypolygelatine and gelatine derivatives or mixtures thereof.
The concentration of the viscosity-increasing sub~tance or substances in the ~olution may be from 0.5 to 50 % by weight, the maximum concentration depending also upon the dissolved substance. Thus, for example, glucose can be used in a concentration of fro~ O.S to 50 ~ by ~eight whereas gelatine ha~ a preferred concentration of from 0.5 to 2 % by weight.
Oxypolygelatine i~ prefera~ly used in a concentration 5'~'~
V g of from 0.5 to 10 % ~y weight.
It i3 also possible to use tensides that have a qimultaneous viscosity-increasing action such, for exam~le, a~ polyoxyethylenepolyoxypropylene polymers having a molecular weight of from 47~0 to 16250.
In thi~ case the concentration of tensides having a viscosity-increasing action is from 1 to 20 %, prefer-ably from 3 to 10 % by wei~ht. The tenside or tenside mixture is preferably dissolved in the liquid carrier in the presence of the viscosity-increasing substance or substances, As the aqueous liquid carrier there may be used water itself or an aqueous solution ~ontaining a water-miscible liquid. There may he mentioned, more especially~
aqueous solutions that are physiologically tolerable, such, for example, as physiological electrolyte solutions such, for example, as physiological common salt solution, Ringer solution or aqueous solutions of sodium chloride, calcium chloride, sodium bicarbonate, sodium citrate, sodium acetate or sodium tartrate, or salt solutions such, for example, as those customarily used as infusion solutions. It will be understood that the solution may be, for example, a colloidal solution.
In a preferred embodiment of the invention the liquid medium used in maXing up the ultrasonic contrast agent comprises a sterile aqueous solution containing 10 % by weiyht of lactose and 0~5 % by weight of a polyoxyethylenepolyoxypropylene polymer having a 5'i"~

molecular weight of from 6800 to 8975 (Pluronic ~ F 68).
Th~ solution may be plac~d in a multi-vial and shaken vigorously with air to produce a dispersion of micro-bubbles.
Thus, for example, for the ultrasonic contrast image of the right ventricle of a dog (beagle of 1702 kg bvdy weight, 2.5 years old, male, closed thorax), OD 3 ml of a micro-bubble Ruspension is used that has been produced by vigorously swirling 2 ml of 5 % by weight strength aqueous sterilised Pluronic ~ F 6a solution and 8 ml of aqueous sterilised 5 ~ strength by weight of glucose solution in a sterile air atmosphere.
To produce the swirling action, the Pluronic ~ F 68 solution was drawn up using an injection syringe, this solution was sprayed into a multi-vial containing the glucose solution at a high discharge speed and then shaken. The 3ize distribution of the micro-bubbles was determined 2 minutes after manufacture using a Cilas granulometer 715 and was 35 ~m for 50 % of the micro-bubbles. The visualisation and recoxding of theultrasonic echoes and their diagnostic evaluation may be efected in a manner known per se and is deseribed, for example, in U.S. Patent No. 4 276 885 and by H. LD Wyatt et al~ in Circulation 60, page 1134 ff -(1979).
The present invention also provides a method of ultrasonic diagnosis of the human or animal body wherein a contrast agent of the invention containing a disper-~ion of micro-bubbles is injected into a part of the human or animal body, especially intravaqcularly, and an ultrasonic image of the micro-bubbles at a slte in the body which it is desired to investigate i9 obtained~
The in~ention still further provides a diagnostj.c kit for use in the ultrasonic diagnoais of the human or animal body, which comprises either as a single solution or separate solutions (A) one or more tensides in an aqueous liquid carrier, and (B) one or more viscosity-increasing substances in an aqueous liquid carrier, the solution or solutions (A) and (B) being in a form (for example, one contained in a syringe and the other in an ampoule or ampoules~ such that on mixing them together the resulting solution will be capable of taking up a plurality of micro-bubbles containing a physiologically tolerable gas, which bubbles will be dispersed in the solutionO Advantageously the kit is a hand-held packO
In one aspect of the invention the kit may comprise a pack containing the solution or solution~ (A) and (B) together with instructions for mixing them together or treating the solution of (A) and (B~ in such manner as to produce a plurality of micro-bubbles containing a physiologically tolerable gas dispersed in the resulting mixture and which is suitable ~or use as an ultrsonic contra~t agent.

- 12 - _ The following Examples illu~trate the invention, the part3 and percentages being by weight unle~s otherwise indicated.
Example 1 .
U~ing an injection ayringe, 2 ml are taXen from a 20 % strength aqueou~ and sterile Pluronic ~ F 68 solution and are ~prayed at the highest practicable discharge speed into 8 ml of an aqueous and sterile 5 % strength glucose ~olution in a sterile 25 ml multi-vial under an atmosphere of air. The mixture i3 then shaken vigorously and the size distribution of the ga~ bub~les i~ measured using a modified Cilas granulometer 715, 2 Minutes after manufacture 50 % of the ga~ bubbles in the suspension had a diameter of < 50 ~m.
The bubble ~uspension so obtained contain~ 4 %
of Pluronic ~ ~ 68 and 4 % of glucose~

E~ample 2 In a manner analogou~ to that described in 20 Example 1 but using a 10 % ~trength aqueous and sterile Pluronic~F 68 solution there is obtained, after the mixing operation, a gas bubble suspen~ion that contains 2 % of Pluronic ~ F 68 and 4 % of glucose.
The 3ize di~tribution of the ga~ bubble3 determined 25 u~ing a Cilas granulometer, i~ ~45 ~m for 50 ~.

jt~yJ
- 13 - ~

Example 3 _ In a manner analogou~ to that described in Example 1 but using a 5 % qtrength aqueous and ~terile Pluronic ~ F 6B ~olution there i3 obtained, after mixing, a ga~ bubble ~u~pen~ion that contains 1 % of Pluronic ~ F 68 and 4 % of glucose.
The bubble si~e, determined u~ing a Cila~
granulometer, i~ < 55 ~m for 50 %.

Example 4 Using an in~ection ~yringe, 2 ml are taken from a 10 % strength aqueous and ~terile Pluronic ~ F 127 solution and are ~prayed at the highest practicable discharge ~peed into 8 ml of a sterile phy~iological common ~alt solution in a sterile 25 ml multi-vial under an atmosphere of air. The mixture i~ then shaken Yigorously. The gas bubble suspension so obtained contain~ 2 ~ of Pluronic ~ F 127 and 0.9 %
. of common ~alt.
The average bubble ~i~e, determined 2 minutes after manufacture u~ing a modified Cilas granulometer, i~ <45 ~m for 50 %.

Example 5 Using an inje~tion ~yringe, 2 ml are taken from an aqueous and sterile 5 % strength Pluronic ~ F 68 solution from which the dissolved air ha~ previously been displaced by argon and are sprayed at the highe~t practicable di~charge ~peed into 8 ml of a sterile 35'î~

aqueous 6 % strength dextran 40 Qolution, from which the di~olved air ha~ al~o keen di~placed by argon, in a ~terile 25/ml multi-vial under an argon atmosphere.
The mixture i~ ~hen sha~en vigorously.
The ~i~e distribution of the argon gas bubble3, determined 2 minute3 after manufacture using a Cila~
granulometer 715, wa4 ~ 55 ~ for 50 %0 '~he resulting gas bubble ~uspension contain~ 1 % of Pluronic ~ F 68 and 4.8 % of dextran 40.

10 Example 6 __ In a manner analcgou~ to that de3cribed in Example 5 but using he7ium Atead ~f argon there i~
obtained, after the mi~ing operati.on, a helium gas bubble ~uspen~ion tha~ ^~ntain~ 1 % of Pluronic ~ F 68 1S and 4~8 % of dextran 4~.
The size di~trib~ion ~ minutes after manufacture ~as ~65 ~m for 50 % o~ the helium gas bubbles.

Example 7 In a manner analc~ous to that de~cribed in 20 Example S but u~ing oxy~en instead of argon there i~
obtained, after the mix~ng operation, an oxygen ga~
bubble 3u~pen~ion that ~ontains 1 % of Pluronic ~ F 68 and 4.8 % of dextran 40_ ~he ~i~e di~tribu .ion 2 minute3 after manu-25 factur~ wa~ <60 ~ for 50 % of the oxygen gas bubble~.

Example 8 _ Using an injection needle, 2 ml are taken from a sterile 5 % ~trength Pluronic ~ F 68 solution in 0.4~ tartaric acid and are ~prayed at the highe~t ~ 5 practicable discharge speed into 8 ml o a sterile 5 % strength glucose solution in 0.1N sodium bicarbonate in a sterile 25 ml multi vial, and the whole i9 ~haken vigorously. A gas bubble suspension is obtained that contains 1 % of Pluronic ~ F 68 and 4 % of glucose~
The size distribution of the gag bubbles, determined

2 minutes after manufacture using a modified Cilas granulometer, is <45 ~m for 50 %.

Example 9 -10 ml portions of a sterile aqueous solution of 10 % by weight of lactose and 0.5 % by weight of Pluronic ~ F 68 are sealed under sterile conditions in conventional 25 ml multi-vials containing ambient air. Upon vigorous shaking and/or repeatedly with-drawing the solution from and injecting it into the vials, a stable (for at least 2 minutes) of micro-bubbles in the solution is produced.

Claims

THE EMBODIMENTS OF THE INVENTION IN WHICH AN EXCLUSIVE
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:

1. An ultrasonic contrast agent for use in diagnosis of the human or animal body, which comprises a solution in an aqueous liquid carrier of (a) a tenside in an amount of from 0.01 to 10% by weight and (b) a viscosity-increasing substance in an amount from 0.5 to 50% by weight, the solution having dispersed in it a plurality of micro-bubbles containing a physiologically tolerable gas.

2. A contrast agent as claimed in claim 1, which comprises a solution containing from 0.5 to 5% by weight of (a).

3. A contrast agent as claimed in claim 2, wherein the tenside is a non-ionic tenside.

4. A contrast agent as claimed in claim 3, where-in the non-ionic tenside is a polyoxyethylenepolyoxy-propylene polymer.

5. A contrast agent as claimed in claim 4, wherein the polyoxyethylenepolyoxypropylene polymer has a molecular weight of from 6800 to 8975.

6. A contrast agent as claimed in claim 4, wherein the polyoxyethylenepolyoxypropylene polymer has a molecular weight of 16250.

7. A contrast agent as claimed in claim 4,wherein the polyoxyethylenepolyoxypropylene polymer has a molecular weight of 13300.

8. A contrast agent as claimed in claim 4, wherein the non-ionic tenside is a polyoxyethylene fatty acid ester.

9. A contrast agent as claimed in claim 8, wherein the non-ionic tenside is a polyoxyethylene stearate.

10. A contrast agent as claimed in claim 2, wherein the tenside is an ionic tenside.

11. A contrast agent as claimed in claim 10, wherein the ionic tenside is sodium lauryl sulphate.

12. A contrast agent as claimed in claim 11, wherein the ionic tenside is sodium dioctyl sulphosuccinate.

13. A contrast agent as claimed in claim 1, wherein the aqueous liquid carrier is water.

14. A contrast agent as claimed in claim 1, wherein the aqueous liquid carrier contains at least one of a water miscible mono- or poly-hydrocyclic alcohol , a phy-siological electrolyte solution and an infusion solution.

15. A contrast agent as claimed in claim 1, 2 or 3, wherein the aqueous liquid carrier contains physiological sodium chloride solution or Ringer solution.

16. A contrast agent as claimed in claim 1, wherein the viscosity-increasing substance is oxypolygelatine.

17. A contrast agent as claimed in claim 1, wherein the viscosity-increasing substance is glucose.

18. A contrast agent as claimed in claim 1, 2 or 3, wherein the viscosity-increasing substance is gelatine.

19. A contrast agent as claimed in claim 1 or 2, wherein the tenside is a non-ionic tenside that has a simultaneous viscosity-increasing action and, if desired, a separate viscosity-increasing substance (b) is omitted.

20. A contrast agent as claimed in claim 1, 2 or 3, wherein the physiologically tolerable gas is sterile air, carbon dioxide, oxygen, nitrogen, helium or argon or a mixture of any two or more such gases.

21. A contrast agent as claimed in claim 1, 2 or 3, which comprises two or more tensides.

22. A contrast agent as claimed in claim 1, 2 or 3, which comprises two or more viscosity-increasing substances.

23. A liquid medium for use in making up the ultra-sonic contrast agent claimed in claim 1, which comprises (i) a solution of one or more tensides in an amount of from 0.01 to 10% by weight in an aqueous liquid carrier, in admixture or conjunction with (ii) a solution of one or more viscosity-increasing substances in an amount of from 0.5 to 50% by weight in an aqueous liquid carrier.

24. A liquid medium as claimed in claim 23, which contains from 0.5 to 50% by weight of the ten-side(s).

25. A liquid medium as claimed in claim 24, wherein the tenside(s) is/are a tenside(s) as claimed in claim 3 or 4.

26. A liquid medium as claimed in claim 24, wherein the liquid carrier is a liquid as claimed in claim 13 or 14.

27. A liquid medium as claimed in claim 24, wherein the viscosity-increasing substance(s) is/are a sub-stance(s) as claimed in claim 16 or 17.

28. A liquid medium as claimed in claim 24, wherein the tenside is a non-ionic tenside that has a sim-ultaneous viscosity-increasing action.

29. A liquid medium as claimed in claim 23, wherein one of the solutions contains from 0.05 to 5% by weight of a physiologically tolerable carbonic acid salt and the other contains an equivalent amount of a physiologically tolerable acid.

30. A process for the manufacture of an ultrasonic contrast agent for use in ultrasonic diagnosis of the human or animal body, which comprises mixing a solution of a tenside in an aqueous liquid carrier with a solution of a viscosity-increasing substance in an aqueous liquid carrier in such manner that the resulting solution contains dispersed in it a plurality of micro-bubbles containing a physio-logically tolerable gas.

31. A process as claimed in claim 30, wherein one of the solution is i injected at high speed into the other solution and the mixture is shaken vigorously.

32. A process as claimed in claim 30, wherein one of the solution is drawn up in an injection syringe.

33. A process as claimed in claim 30, wherein one of the solutions is contained in a vial or multi-vial.

34. A process as claimed in claim 33, wherein the vial or multi-vial contains air, oxygen, carbon dioxide, nitrogen, helium or argon or a mixture of any two or more such gases.

35. A composition comprising a solution in an aqueous liquid carrier of (a) a tenside or tensides in an amount of from 0.01 to 10% by weight and (b) a viscosity?

increasing substance or substances in an amount of from 0.5 to 50% by weight , the solution having dispersed in it a plurality of micro-bubbles containing a physiologically tolerable gas for use in a method of diagnosis practised on the human or animal body.

36. A composition as claimed in claim 35, which contains from 0.5 to 5% by weight of the tenside(s).

37. A composition as claimed in claim 35, wherein the tenside(s) is/are a tenside(s) as claimed in claim 3 or 4.

38. A composition as claimed in claim 35, wherein the liquid carrier is a liquid as claimed in claim 13 or 14.

39. A composition as claimed in claim 35, wherein the viscosity-increasing substance(s) is/are a substance(s) as claimed in claim 16 or 17.

40. A composition as claimed in claim 35 or 36, wherein the tenside is a non-ionic tenside that has a simultaneous viscosity-increasing action.

41. A composition as claimed in claim 35, wherein one of the solutions contains from 0.05 to 5% by weight of a physiologically tolerable carbonic acid salt and the other contains an equivalent amount of a physiologically tolerable acid.