New monoclonal antibodies for diagnostic purposes

The construction of hybridomas producing monoclonal
antibodies for new, more specific diagnostic kits for the
detection of colon cancer and some air allergens.

Diagnostics is a 16 billion dollar industry worldwide, dominated by antibody-based products (immunodiagnostics) which form 30% of the total market and it is increasing at a rate of 15% per annum. New diagnostic assays have a short lead time to market, usually less than 2 years, and offer a rapid return on investment. Monoclonal antibodies are a basic part of these new diagnostics. The main aim of our proposal is the construction of new hybridomas producing monoclonal antibodies with very specific attributes which will enable low cost and rapid diagnostics or detection of the target antigen. Aim 1: Colon Cancer Diagnosis According to the data published in the 'Cancer Facts and Figures' (1990) CZECHOSLOVAKIA was reported as the country with the highest incidence of colorectal cancer. The survival of patients with this cancer is proportional to the stage of tumor invasion and spread at the time of diagnosis. On the other hand, colorectal carcinoma is considered one of the most curable forms of cancer. Consequently there is considerable interest in identifying tumor-specific markers that would enable the early diagnosis of this cancer. Ideally, the marker would be specific for a particular disease and the detection method sensitive enough to provide an early indication of disease state. For a number of years the diagnosis of colon cancer included testing for faecal occult blood (FOB) using a colorimetric assay. The colorimetric reagent, tetra-methyl- benzidine is still utilised today in many test pads. This diagnostic test is prone to interference due to dietary intake of red meat, vitamin C, etc. Common medications such as aspirin, corticosteroids, anticoagulants also interfere with the test result. In addition, false positive results have been blamed on menstruation, constipation or heamorrhoids. The development of diagnostic test strip utilising a monoclonal antibody against human haemoglobin has relieved some of the previous problems associated with interfering substances in faecal sample. However, the detection of blood in stool does not mean the patient has colon cancer and may be an indication of a number of other gastrointestinal disorders such as fissures and diverticular disease. At present, four biotechnology companies share the market demand in the USA which has been estimated to be worth more than 100,OOO,OOO US dollars per annum. A growing number of researchers are reporting new markers for colon cancer. For example, mucin antigen (Guang YY an Shamsuddin AM. Arch. Pathol. Lab Med. 1995: 119,454), cyclooxygenase-2 (Sheehan KM et al. JAMA, 1999: 282,1254) and CD44 variants (Hsieh HF et al. Mol. Pathol. 1999: 52,25). The aim of this part of our project will be the development of new monoclonal antibodies to some of colon cancer markers (haemoglobin, mucin, CD44 variants, etc.) with the aim to improve both the sensitivity and specificity of diagnostic tests for colon cancer by incorporating reagents designed to detect a marker, which is detectable in both stool and serum samples of patients suffering from colon cancer. Aim 2: Allergen ELISA kit Occurring in industrial fumes and wastes, in diesel exhaust and in incomplete combustion, polycyclic aromatic hydrocarbons (PAHs) and nitro-PAHs are hazardous environmental pollutants to which man is exposed in the environment and in the work atmosphere of several industries such as coke oven plants, the electrochemical industry, impregnation plants, electrode factories, and in various manufacturing plants with combustion processes. In contrast to most of the environmental pollutants currently discussed, the hazardous potential of these compounds is supposed to be rather underestimated. They are metabolized by mixed-function cytochrome-dependent oxygenases that my form metabolites which are either excreted in urine or react as ultimate carcinogens with DNA or other macromolecules. Therefore, some of these compounds exert mutagenicity and carcinogenicity through formation of adducts with DNA and harmful effects on mismatch repair. The INTERNATIONAL AGENCY FOR RESEARCH ON CANCER (IARC) and UNITED STATES' ENVIRONMENTAL PROTECTION AGENCY (USEPA) have classified several of these compounds as probable or possible human carcinogens. Little is known about the human cancer risk of the widespread distribution of low levels of PAHs in food products and in polluted air. In the new European Drinking Water Guideline (98/83/EC) special attention is given to benzo(a)pyrene (BAP) by setting a threshold-limit of 10 ng/L for this single compound. For the analysis of environmental samples, purification and enriching followed by a powerful chromatographic separation and identification is required. Most utilized is HPLC with UV or fluorescent detection. Immunochemical methods are increasingly applied for the determination of environmental analytes. Exhibiting comparable sensitivity, in many cases they have been proven to be less time-consuming and more cost-effective alternatives to traditional chromatographic techniques, because sample preparation can generally be reduced to a great extent or omitted completely. In the past, we have developed an ELISA for PAHs that was used for the detection of these compounds in water and soil samples. The polyclonal antiserum recognizes mainly the lower annelated (3- and 4-ringed aromatic) compounds. Cross-reactivity with BAP is about 15%. Within the EUREKA project a monoclonal antibody should be prepared which exhibits higher binding for BAP. Keywords: monoclonal antibodies, diagnostics, allergens.
Project ID: 
2 334
Start date: 
Project Duration: 
Project costs: 
1 010 000.00€
Technological Area: 
Diagnostics, Diagnosis
Market Area: 

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