US4558007A
Expired / Lapsed
US
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Process for detecting LDHk isozyme activity in human serum for use as a diagnostic aid and for monitoring response to cancer therapy
Assignee: Health Research Inc
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Filing: 1983-06-17
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Granted: 1985-12-10
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Expires: 2003-06-17
Abstract
Abstract An improved assay procedure for quantitatively detecting LDHk isozyme in human serum is provided which utilizes previously known electrophoretic gel separation procedures combined with an improved staining method wherein the staining solution has a constant temperature of about 37 DEG C. and a pH of about 8.0, and is contacted with the gel matrix for at least 3 hours in the absence of oxygen and light. This improved assay procedure may be used as an adjunct to other procedures for diagnosing the presence of primary cancer, as a post-operative indicator of metastatic cancer and in monitoring the success of cancer therapy.
Inventors
- 👤 Garth R. Anderson
- 👤 Kenneth F. Manly
- 👤 Arnold Mittelman
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-30 23:07
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/53; G01N33/575; G01N33/5758; G01N33/57585; C; C12; C12Q; C12Q1/00; C12Q1/26; C12Q1/32; G01N2333/00; G01N2333/90; G01N2333/902; G01N2333/904; G01N2800/00; G01N2800/52
Claims Analysis
Total Claims: 16
Claim 1
Independent Claim
83 words
In a known process for quantitatively detecting LDH k isozyme in human serum utilizing reverse polarity electrophoresis on an imidazole/borate buffered polyacrylamide gel matrix according to known techniques, followed by staining the gel matrix using standard nitroblue tetrazolium/phenazine methasulfate or an equivalent staining solution, the improvement comprising continuously contacting said gel matrix with said staining solution having a constant temperature between 30°-40° C. for at least 3 hours in the absence of both oxygen and light to provide a suitable stain reaction time.
Claim 2
Dependent Claim
83 words
In a known process for quantitatively detecting LDH k isozyme in human serum utilizing reverse polarity electrophoresis on an imidazole/borate buffered polyacrylamide gel matrix according to known techniques, followed by staining the gel matrix using standard nitroblue tetrazolium/phenazine methasulfate or an equivalent staining solution, the improvement comprising continuously contacting said gel matrix with said staining solution having a constant temperature between 30°-40° C. for at least 3 hours in the absence of both oxygen and light to provide a suitable stain reaction time.
Claim 3
Dependent Claim
16 words
The improved process according to claim 1 wherein said gel is submerged within the staining solution.
Claim 4
Dependent Claim
22 words
The improved process according to claim 2 wherein the pH of the staining solution is maintained between 7.8-9.0 during said reaction time.
Claim 5
Dependent Claim
24 words
The improved process according to claim 3 wherein the temperature of the staining solution is maintained at about 37° C. during said reaction time.
Claim 6
Dependent Claim
21 words
The improved process according to claim 4 wherein the pH of the staining solution is about 8.0 during said reaction time.
Claim 7
Dependent Claim
29 words
The improved process according to claim 5 wherein the staining solution used during said reaction time has a pH of about 8.0 and a temperature of about 37° C.
Claim 8
Dependent Claim
15 words
The improved process according to claim 6 wherein said reaction time is between 3-8 hours.
Claim 9
Dependent Claim
16 words
The improved process according to claim 7 wherein said reaction time is at least 4.5 hours.
Claim 10
Dependent Claim
17 words
The improved process according to claim 8 wherein said staining process is performed in a nitrogen environment.
Claim 11
Dependent Claim
67 words
A method for determining in a patient the presence or absence of cancer which comprises extracting from said patient a sample of blood, and measuring the quantitative relative activity level of LDH k isozyme present within the serum of said sample by the method of claim 1 and comparing said level to the level of LDH k measured in the serum of an individual free of cancer.
Claim 12
Dependent Claim
30 words
The method of claim 10 wherein the patient's level of LDH k is compared with a standard curve showing average nomal or base line levels of serum LDH k activity.
Claim 13
Dependent Claim
50 words
The method of claim 11 wherein the LDH k activity is determined in said patient's serum after the patient has undergone treatment for cancer and compared with the level of LDH k activity in the serum of an individual free of cancer to determine whether metastasis has occurred after therapy.
Claim 14
Dependent Claim
48 words
The method of claim 12 wherein the LDH k activity level is determined in said patient's serum after the patient has undergone treatment for cancer and compared with the level of LDH k activity in the serum of an individual free of cancer to monitor cancer treatment response.
Claim 15
Dependent Claim
36 words
A method using LDH k isozyme as a serum marker for detecting the presence of cancer in a patient wherein the serum level of the patient's LDH k is determined by the method of claim 1.
Claim 16
Dependent Claim
117 words
A diagnostic test kit for performing the method of claim 1 comprising: (a) a given quantity of a suitable electrophoresis matrix for use in electrophoretically separating LDH k isozyme; (b) A given quantity of a borate buffer having a concentration greater than or equal to 10mM borate ion for treating the serum prior to or simultaneously with electrophoresis; (c) a given amount of appropriate staining reagents for the colorimetric determination of LDH k ; (d) means for excluding light and oxygen from the testing environment; (e) a positive control comprising an aliquot of purified LDH k ; (f) directions for the performance of said method; and (g) a container for housing the materials present in steps (a)-(f).
References Cited (Backward Citations)
US4258131A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1988-12-09 | 1989-06-10 | 1989-12-10 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1992-12-09 | 1993-06-10 | 1993-12-10 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1996-12-09 | 1997-06-10 | 1997-12-10 | $7700.00 | Window Closed / Lapsed |