Method for quantitation of collagen in tissue
원문보기
IPC분류정보
국가/구분
United States(US) Patent
등록
국제특허분류(IPC7판)
C12Q-001/00
G01N-033/48
출원번호
US-0858737
(2007-09-20)
등록번호
US-7491541
(2009-02-17)
발명자
/ 주소
Villanueva,Patricia A.
McNulty,Amy K.
Beniker,Herbert D.
Kieswetter,Kristine
출원인 / 주소
KCI Licensing, Inc.
인용정보
피인용 횟수 :
2인용 특허 :
85
초록▼
For successful wound healing to occur, the newly formed skin must generate tensile strength through collagen deposition. Measurement of collagen in the granulating wound bed may be predictive of successful wound healing. Existing methods for collagen measurement either require specialized equipment,
For successful wound healing to occur, the newly formed skin must generate tensile strength through collagen deposition. Measurement of collagen in the granulating wound bed may be predictive of successful wound healing. Existing methods for collagen measurement either require specialized equipment, or do not allow for discrimination of potential interfering molecules. Herein described is an accurate, specific and reliable method to extract and quantify collagen from tissue utilizing high pressure liquid chromatography techniques. The method is sensitive enough to measure the small amounts of collagen found in newly healing wounds.
대표청구항▼
What is claimed is: 1. A method of assessing wound healing progress comprising: (a) obtaining a tissue sample from a wound site of a subject; (b) processing the tissue sample by a method comprising: (i) removing fat from the tissue sample; (ii) dehydrating the tissue sample; (iii) hydrolyzing the t
What is claimed is: 1. A method of assessing wound healing progress comprising: (a) obtaining a tissue sample from a wound site of a subject; (b) processing the tissue sample by a method comprising: (i) removing fat from the tissue sample; (ii) dehydrating the tissue sample; (iii) hydrolyzing the tissue sample to amino acids; and (iv) derivatizating the tissue sample; (c) separating glycine, proline, and hydroxyproline in the processed tissue sample by high-pressure liquid chromatography to obtain analyte peaks areas; (d) determining concentrations of the glycine, proline, and hydroxyproline in the tissue; (e) correlating the concentrations of the glycine, proline, and hydroxyproline in the tissue with an amount of total collagen in the tissue; and (f) comparing the amount of total collagen in the tissue at the wound site with the amount of collagen in non-wounded tissue to assess wound healing progress. 2. The method of claim 1, wherein the tissue is skin. 3. The method of claim 1, wherein the tissue is granulated. 4. The method of claim 1, wherein steps (a)-(f) are repeated at least once, and the tissue sample is obtained from the same wound site of the same subject for each repetition. 5. The method of claim 4, wherein the steps (a)-(f) are repeated at least once during a time period of between about 3 days and about 2 years. 6. The method of claim 4, further comprising comparing the amount of total collagen in the tissue at a first repetition with the amount of total collagen in the tissue at one or more subsequent repetitions. 7. The method of claim 1, wherein the subject is a mammal. 8. The method of claim 7, wherein the mammal is a human. 9. The method of claim 1, wherein the high-pressure liquid chromatography is performed on a C18 column. 10. The method of claim 9, wherein the C18 column has a pH range of about 1 to about 12. 11. The method of claim 1, wherein elution of the glycine, proline, and hydroxyproline during the high-pressure liquid chromatography is measured at 436 nm with a photodiode array detector. 12. The method of claim 1, wherein the high-pressure liquid chromatography is reversed phase high-pressure liquid chromatography. 13. The method of claim 1, wherein the high-pressure liquid chromatography has a mobile phase of 70% 25 mM potassium phosphate, pH 11.0 buffer, and 30% acetonitrile. 14. The method of claim 1, wherein determining the concentrations of the glycine, proline, and hydroxyproline in the tissue comprises: (a) obtaining samples with known concentrations of glycine, proline, and hydroxyproline; (b) separating the glycine, proline, and hydroxyproline in the samples by high-pressure liquid chromatography; (c) plotting the known concentrations of glycine, proline, and hydroxyproline on a graph x-axis by analyte peak areas on the graph y-axis to devise a linear standard curve; and (d) calculating the concentrations of each of the glycine, proline, and hydroxyproline in the processed tissue sample using a formula: wherein b is the y-intercept and m is the slope of the linear standard curve; and (e) calculating the concentrations of each of the glycine, proline, and hydroxyproline in the tissue using a formula: 15. The method of claim 1, wherein correlating the concentrations of the glycine, proline, and hydroxyproline in the tissue with the amount of total collagen in the tissue comprises using a formula: wherein C is the sum of the percent composition of glycine, proline, and hydroxyproline in collagen divided by 100. 16. The method of claim 15, wherein C equals 0.55.
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이 특허에 인용된 특허 (85)
Ward William J. (Hull GBX) Shorthouse Joanne (Hull GBX), Adhesive dressing for retaining a cannula on the skin.
Errede Louis A. (North Oaks MN) Stoesz James D. (St. Paul MN) Winter ; deceased George D. (late of St. Paul MN by Jenny Upton ; personal representative), Composite wound dressing.
Todd Robert J. (Salt Lake City UT) Yagge Jaime E. (Salt Lake City UT) Lowe James E. (Durham NC) Wonder Terry M. (Salt Lake City UT), Medical suction apparatus.
Ferdman Ariel (12 Hillside Ave. Melrose MA 02176) Kuo Jing-wen (Boxboro MA) Miller David (Brookline MA) Pinsky Vladimir (Brighton MA) Richards William D. (Medway MA) Swann David (Cambridge MA), Method and device for wound closure.
Moriuchi Yousuke (Fujinomiya JPX) Ishida Toshinobu (Fujinomiya JPX) Kousai Tadashi (Fujinomiya JPX), Method of securing a catheter body to a human skin surface.
Luheshi Abdul B. N. (Hull GB3) Smalley Robert K. (Urmston GB3) Kennewell Peter D. (Okus GB3) Westwood Robert (Kingston Bagpuize GB3), Process for the preparation of azabicyclo compounds.
Richmond James W. (Kalamazoo MI) Tice Robert G. (Portage MI) Booth ; III William M. (Paw Paw MI), Vacuum wound drainage system and lipids baffle therefor.
McNeil Charles B. (5960 Arbour Ave. Edina MN 55436) McEvoy Thomas J. (13103 Baker Trail Minnetonka MN 55343), Wearable, variable rate suction/collection device.
Lock Peter M. (327 Lordswood La. Petrosa ; Walderslade ; Chatham ; Kent GB2) Webb David R. (9 Lambourn Way Lordwood ; Chatham ; Kent GB2), Wound dressing materials.
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