Method of detecting the presence of CD155 for diagnosis of cancer and to determine treatment
AssigneeThe Research Foundation of State University of New York
InventorsMatthias Gromeier, Eckard Wimmer
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Abstract
The present invention relates to a method of diagnosing, classifying and grading of tumor growths and to determine whether the use of chimeric polioviruses is a proper course for the treatment of the tumors. More particularly, the method is directed to the use of antibodies to a poliovirus receptor (PVR), CD155, to detect the presence of CD155 on tumor cells in various organs, such as: breast, colon, bronchial passage, epithelial lining of the gastrointestinal, upper respiratory and genito-urinary tracts, liver, prostate and the brain.
Claims13
A method for detecting CD155 in tumor tissue comprising the steps:
- a) treating a surgical explant tumor tissue to prepare a thin section of the tumor tissue for mounting the section on a pre-coated microscopic slide;
- b) fixing and blocking the section on the slide;
- c) reacting the section with an anti-CD155 monoclonal antibody to bind CD155;
- d) reacting a secondary antibody conjugated to a chromophore with the bound anti-CD155 to form a complex;
- e) developing a color by reacting the complex with a chromogenic substrate for the chromophore;
- and
- f) detecting the presence of color which indicates the presence of CD155.
A method for detecting CD155 in tumor tissue by an immunoassay comprising the steps:
- a) homogenizing 50-500 mg of surgical explant tumor tissue in about 200 μl of a solubilization buffer comprising PBS containing 0.5% nonylphenoxy polyethoxy ethanol (SP40™) for 10 min. on ice to completely disintegrate the tumor tissue;
- b) centrifuging the homogenized tumor tissue to separate out the supernatant fluid;
- c) detecting the presence of CD155 in the supernatant fluid in an immunodot assay, an immunoprecipitation assay, an immunoblot assay or an ELISA procedure wherein the binding of a monoclonal anti-CD155 antibody to the tumor tissue indicates the presence of CD155 in the tumor tissue.
A method for detecting CD155 in tumor tissue comprising the steps:
- a) treating tumor tissue immediately after surgical removal by one of the following steps:
- i) shock-freezing the tumor tissue immediately by one of the following steps:
- immersing in cold isopentane cooled on dry ice, immersing in liquid nitrogen, immersing in powdered dry ice, or submerging in a freezing compound comprising a water soluble glycol and resins matrix for cryostat sectioning, followed by placing into powdered dry ice;
- or
- ii) fixing the sample by using an appropriate fixing solution selected from the group consisting of:
- 2% neutral buffered paraformaldehyde, 2% neutral buffered paraformaldehyde/0.2% glutaraldehyde, ice cold acetone, 25% glacial acetic acid in 100% ethanol;
- or
- iii) freezing the fixed sample by immersion in 30% sucrose in phosphate buffered saline over night and placing in a freezing compound comprising a water soluble glycol and resins matrix for cryostat sectioning at −70° C. for at least 15 min.;
- b) sectioning the tumor tissue on a cryostat into a thickness of about 5-25 μm;
- c) placing and fixing the thin sections on silane treated microscopic slides for immunohistochemical assay using as a primary antibody a monoclonal anti-CD155 antibody for detecting the presence of CD155 in the tumor tissue;
- d) developing and detecting a chromogenic response by:
- i) immersing the sections for 5 min. in acetate buffer comprising:
- 0.01M acetic acid, 0.035M sodium acetate trihydrate, at pH 5.4,
- ii) developing the color by immersing the sections in an acetate buffer containing a substrate for horseradish peroxidase and 0.6% hydrogen peroxide,
- iii) rinsing the sections in PBS and distilled water and drying and mounting the rinsed sections with an appropriate mounting medium, and
- e) detecting the presence of color in the tumor tissue which indicates the presence of CD155.
Dependencies are inferred from the claim text.