Antibody hybridization

Western blot and immunohistochemical detection are the two most widely used techniques to confirm the expression of a given P450 isoform in isolated brain regions or thin slices. Antibodies detect actual protein, 

Cytochrome P450 pathways in human brain cholesterol, steroid hormones, vitamin D, and bile 

17-hydroxy- 3[3HSD-1/2 lyp.hydroxy- CYP IA 11-Deoxycortisol pregnenolone progesterone 

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Concentrations of oligonucleotide probes and antibody, hybridization temperature, and incubation times correspond to those used in the authors laboratory. However, it may be necessary to optimize each of these parameters, with those given in the protocol serving as a good starting point. For oligonucleotide probes, the authors have found 1 pmol of each as a universally suitable concentration. 

Youle R, Neville D. Kinetics of protein synthesis inactivation by ricin-anti-thy.1.1 monoclonal antibody hybrids Role of the ricin B subunit demonstrated by reconstitution. J Biol Chem. 1982 267 1598-1601. 

Houghton AN, Brooks H, Cote RJ, Taormina MC, Oettgen HF, Old LJ (1983) Detection of cell surface and intracellular antigens by human monoclonal antibodies. Hybrid cell lines derivedfrom lymphocytes of patients with malignant melanoma. J Exp Med 158 53-65... 

As the result of high specificity and sensitivity, nucleic acid probes are in direct competition with immunoassay for the analytes of some types of clinical analytes, such as infectious disease testing. Assays are being developed, however, that combine both probe and immunoassay technology. In such hybrid probe—immunoassays, the immunoassay portion detects and amplifies the specific binding of the probe to a nucleic acid. Either the probe per se or probe labeled with a specific compound is detected by the antibody, which in turn is labeled with an enzyme or fluorophore that serves as the basis for detection. 

Mammalian Cells Unlike microbial cells, mammalian cells do not continue to reproduce forever. Cancerous cells have lost this natural timing that leads to death after a few dozen generations and continue to multiply indefinitely. Hybridoma cells from the fusion of two mammalian lymphoid cells, one cancerous and the other normal, are important for mammalian cell culture. They produce monoclonal antibodies for research, for affinity methods for biological separations, and for analyses used in the diagnosis and treatment of some diseases. However, the frequency of fusion is low. If the unfused cells are not killed, the myelomas 1 overgrow the hybrid cells. The myelomas can be isolated when there is a defect in their production of enzymes involved in nucleotide synthesis. Mammahan cells can produce the necessary enzymes and thus so can the fused cells. When the cells are placed in a medium in which the enzymes are necessaiy for survival, the myelomas will not survive. The unfused normal cells will die because of their limited life span. Thus, after a period of time, the hybridomas will be the only cells left ahve. 

Humanized Monoclonal Antibodies Humoral Immunity HVA Ca2+ Channels Hybridization... 

Built-in functional specific recognition and binding, e.g., antigen binding by antibodies or hybridization of nucleic acids ... 

Linardos, T., "Kinetics of Monoclonal Antibody Production in Chemostat Hybrid-oma Cultures", Ph D. thesis, Dept, of Chemical Petroleum Engineering, University of Calgary, AB, Canada, 1991. 

VI3. Vora, S and Francke, U., Assignment of the human gene for liver-type 6-phosphofructokinase isozyme (PFKL) to chromosome 21 by using somatic cell hybrids and monoclonal anti-L antibody. Proc. Natl. Acad. Sci. U.S.A. 78,3738-3742 (1981). 

In the Hybrid-Capture assay (Digene), a full-length RNA probe is hybridized to denatured HBV DNA in solution and the hybrids are captured on the surface of a tube coated with anti DNA RNA hybrid antibody. The bound hybrids are reacted with antihybrid antibody labeled with alkaline phosphatase. A chemiluminescent substrate is converted to a luminescent compound by the bound alkaline phosphatase. Light emission is measured in a luminometer and the concentration of HBV DNA, in pg/ml, is determined from a standard curve. The concentrations of the standards are determined spectrometrically (A260nm/A280nm). 

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