You’re using a public version of DrugPatentWatch with 5 free searches available | Register to unlock more free searches. CREATE FREE ACCOUNT

Last Updated: March 28, 2024

Claims for Patent: 5,753,485


✉ Email this page to a colleague

« Back to Dashboard


Summary for Patent: 5,753,485
Title: Purified mixture of collagenase I, collagenase II and two other proteases
Abstract:A purified enzyme mixture useful for isolating cells or cell clusters from tissue is disclosed. The mixture includes at least two collagenase enzymes, at least two other proteases, and additional non-protease components. The mixture is purified by removing at least some of the non-protease components. The purified mixture may then be used to isolate cells or cell clusters from tissue. Also disclosed are the essential components of the purified enzyme mixture as well as preferred ranges and ratios of these essential components for isolating cells or cell clusters from tissue. Finally, a dissociation system is disclosed that can be used with the purified enzyme mixture to dissociate tissue and recover cells or cell clusters.
Inventor(s): Dwulet; Francis E. (Greenwood, IN), Ellis; Bernice B. (Greenfield, IN), Gill; John F. (Indianapolis, IN), Jacobsen; Linda B. (Indianapolis, IN), Smith; Marilyn E. (McCordsville, IN), Waters; David G. (Indianapolis, IN)
Assignee: Boehringer Mannheim Corporation (Indianapolis, IN)
Application Number:08/590,957
Patent Claims:1. An enzyme composition prepared by combining defined amounts of proteases said composition comprising collagenase I and collagenase II from C. histolyticum, having an A.sub.280 /A.sub.360 ratio of greater than 30 and two other proteases, wherein the ratio of the mass of collagenase II to the mass of collagenase I plus the mass of the collagenase II in the composition is about 0.3 to about 0.6.

2. The composition of claim 4 wherein the ratio of the mass of collagenase II to the mass of collagenase I plus the mass of collagenase II is about 0.4 to about 0.45.

3. The composition of claim 1 wherein the collagenase I and collagenase II have an A.sub.280 /A.sub.360 ratio greater than 100.

4. The enzyme composition of claim 1 wherein at least one of the two other proteases is a neutral protease.

5. The enzyme position of claim 4 wherein the neutral protease is selected from the group consisting of thermolysin, dispase and C. histolyticun neutral protease.

6. The composition of claim 5 wherein the ratio of the mass of collagenase II to the mass of collagenase I plus the mass of collagenase II is about 0.4 to about 0.45.

7. The enzyme composition of claim 4 wherein the neutral protease is thermolysin.

8. The enzyme composition of claim 7 wherein the ratio of the total FITC casein activity of the enzyme composition to the total Wunsch units of activity of the masses of collagenase I and collagenase II in the composition is about 19:1 to about 70:1.

9. The enzyme composition of claim 4 wherein the neutral protease is dispase.

10. The enzyme composition of claim 9 wherein the ratio of the total FITC casein activity of the enzyme composition to the total Wunsch units of activity of the masses of collagenase I and collagenase II in the composition is about 19:1 to about 62:1.

11. The enzyme composition of claim 4 wherein the neutral protease is dispase or thermolysin and the ratio of total FITC casein activity of the enzyme composition to the total Wunsch units of the masses of collagenase I and collagenase II in the composition is about 41:1.

12. The enzyme composition of claim 4 wherein the two other proteases are a neutral protease and clostripain.

13. The enzyme composition of claim 12 wherein the neutral protease is selected from the group consisting of thermolysin, dispase and C. histolyticum neutral protease.

14. The enzyme composition of claim 12 wherein the neutral protease is thermolysin.

15. The enzyme composition of claim 14 wherein the ratio of the total FITC casein activity of the enzyme composition to the total Wunsch units of activity of the masses of collagenase I and collagenase II in the composition is about 19:1 to about 70:1.

16. The enzyme composition of claim 12 wherein the neutral protease is dispase.

17. The enzyme composition of claim 16 wherein the ratio of the total FITC casein activity of the enzyme composition to the total Wunsch units of activity of the masses of collagenase I and collagenase II in the composition is about 19:1 to about 62:1.

18. The enzyme composition of claim 12 wherein the neutral protease is dispase or thermolysin and the ratio of total FITC casein activity of the enzyme composition to the total Wunsch units of the masses collagenase I and collagenase II in the composition is about 41:1.

19. The composition of claim 12 wherein the ratio of the mass of collagenase II to the mass of collagenase I plus the mass of collagenase II is about 0.4 to about 0.45.

20. A method of preparing an enzyme composition adapted for isolating living cells from tissue, said method comprising the steps of combining collagenase I, collagenase II, and clostripain from C. histolyticum wherein the ratio of the mass of collagenase II to the mass of collagenase I plus the mass of collagenase II is about 0.3 to about 0.6; and

adding an amount of a neutral protease sufficient to raise the total FITC casein activity of the enzyme composition to a level such that the ratio of the total FITC casein activity of the enzyme composition to the total Wunsch unit activity of the masses of collagenase I and collagenase II in the enzyme composition is about 19:1 to about 70:1.

21. The method of claim 20 wherein the ratio of the mass of collagenase II to the mass of collagenase I plus the mass of collagenase II is about 0.4 to about 0.45.

Make Better Decisions: Try a trial or see plans & pricing

Drugs may be covered by multiple patents or regulatory protections. All trademarks and applicant names are the property of their respective owners or licensors. Although great care is taken in the proper and correct provision of this service, thinkBiotech LLC does not accept any responsibility for possible consequences of errors or omissions in the provided data. The data presented herein is for information purposes only. There is no warranty that the data contained herein is error free. thinkBiotech performs no independent verification of facts as provided by public sources nor are attempts made to provide legal or investing advice. Any reliance on data provided herein is done solely at the discretion of the user. Users of this service are advised to seek professional advice and independent confirmation before considering acting on any of the provided information. thinkBiotech LLC reserves the right to amend, extend or withdraw any part or all of the offered service without notice.