Patent US4338242A successfully refreshed from Google Patents.
Patent US4279774A successfully refreshed from Google Patents.
US4279774A
Expired / Lapsed
US
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Method of removing lipid inhibitors from limulus amebocyte lysate
Assignee: Lonza Walkersville Inc
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Filing: 1979-11-13
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Granted: 1981-07-21
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Expires: 1999-11-13
Abstract
Abstract The improved method of the present invention involves the removal of lipid inhibitors from Limulus amebocyte lysate to increase the sensitivity and quality of the lysate. The method comprises intimately contacting, as by mixing and stirring, Limulus amebocyte lysate with a selected binary liquid solvent system for a time sufficient to draw water into the solvent system from the lysate and to effectively extract and denature the lipid inhibitors in the lysate. The ability of the binary solvent to draw in water from the lysate and form, in effect, a new tertiary system is the important factor in this methodology. The amount of water drawn into the solvent is controlled by the amount of polar solvent in the solvent system. After this extraction and denaturation, the lysate is then separated from the solvent system and recovered in purified form of increased sensitivity.
Inventors
- 👤 Gene K. Lindsay
- 👤 Andrew J. O'Beirne
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-09 04:46
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/579
Claims Analysis
Total Claims: 16
Claim 1
Independent Claim
160 words
An improved method of removing lipid inhibitors from Limulus amebocyte lysate to increase the sensitivity and quality of the lysate, said method comprising: (a) intimately contacting Limulus amebocyte lysate with a selected binary liquid solvent system containing first and second selected organic solvents of differing polarities, said first solvent being of low polarity registering not in excess of about 0.5 on the elutotrophic series scale, said second solvent being of high polarity registering at least about 0.8 on said elutotrophic series scale, said second solvent being present in said solvent system in a concentration of about 2-25% by volume of said system, said solvents being present in an amount and contacting said lysate for a time sufficient to draw water into said solvent system from said lysate and extract and denature lipid inhibitors in said lysate; and, (b) separating said Limulus amebocyte lysate and said solvent system from each other and recovering said lysate in purified form having improved sensitivity.
Claim 2
Dependent Claim
160 words
An improved method of removing lipid inhibitors from Limulus amebocyte lysate to increase the sensitivity and quality of the lysate, said method comprising: (a) intimately contacting Limulus amebocyte lysate with a selected binary liquid solvent system containing first and second selected organic solvents of differing polarities, said first solvent being of low polarity registering not in excess of about 0.5 on the elutotrophic series scale, said second solvent being of high polarity registering at least about 0.8 on said elutotrophic series scale, said second solvent being present in said solvent system in a concentration of about 2-25% by volume of said system, said solvents being present in an amount and contacting said lysate for a time sufficient to draw water into said solvent system from said lysate and extract and denature lipid inhibitors in said lysate; and, (b) separating said Limulus amebocyte lysate and said solvent system from each other and recovering said lysate in purified form having improved sensitivity.
Claim 3
Dependent Claim
21 words
The improved method of claim 1 wherein said system is used in a volume ratio of about 1:2 to said lysate.
Claim 4
Dependent Claim
23 words
The improved method of claim 1 wherein said first solvent is an aromatic and said second solvent is a low molecular weight alcohol.
Claim 5
Dependent Claim
21 words
The improved method of claim 3 wherein said system comprises a major concentration of benzene and a minor concentration of methanol.
Claim 6
Dependent Claim
22 words
The improved method of claim 4 wherein said methanol is in a concentration of about 2-5 volume per cent in said system.
Claim 7
Dependent Claim
13 words
The improved method of claim 3 wherein said system comprises benzene and ethanol.
Claim 8
Dependent Claim
22 words
The improved method of claim 6 wherein said ethanol is present in said system in a concentration of about 2-5 volume percent.
Claim 9
Dependent Claim
17 words
The improved method of claim 3 wherein said system comprises toluene and a low molecular weight alcohol.
Claim 10
Dependent Claim
22 words
The improved method of claim 8 wherein said alcohol comprises methanol in a concentration of about 2-5 volume percent in said system.
Claim 11
Dependent Claim
22 words
The improved method of claim 8 wherein said alcohol comprises ethanol in a concentration of about 2-5 volume percent in said system.
Claim 12
Dependent Claim
19 words
The improved method of claim 1 wherein said system comprises a multi-halogenated paraffin and a low molecular weight alcohol.
Claim 13
Dependent Claim
26 words
The improved method of claim 11 wherein said system comprises trichloroethane and ethanol, the latter in a concentration of about 2-5 volume percent in said system.
Claim 14
Dependent Claim
26 words
The improved method of claim 11 wherein said system comprises trichloroethane and propanol, the latter in a concentration of about 2-5 volume percent in said system.
Claim 15
Dependent Claim
40 words
The improved method of claim 1 wherein said contacting is effected by mixing said lysate with said system for approximately one-half hour, and wherein said separating is effected by decanting said system from said lysate, whereupon said lysate is lyophilized.
Claim 16
Dependent Claim
43 words
The improved method of claim 1 wherein said first solvent is selected from the group consisting of benzene, trichloroethane and toluene, and mixtures thereof, and wherein said second solvent is selected from the group consisting of methanol, ethanol and propanol, and mixtures thereof.
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USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1984-07-20 | 1985-01-19 | 1985-07-21 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1988-07-20 | 1989-01-19 | 1989-07-21 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1992-07-20 | 1993-01-19 | 1993-07-21 | $7700.00 | Window Closed / Lapsed |