Radiolabeled probes

Mycobacteria of the Mycobacterium avium complex are implicated in disseminated bacterial infections in AIDS patients. RFLP studies followed by hybridization with radiolabeled probe specific for an insertion sequence in M. avium (IS 1311) have been useful for typing M. avium stains (R2). A variety of molecular techniques are available for the diagnosis of Chlamydia trachomatis infection. In addition to PCR, a method based on the ligase chain reaction has also been found to be sensitive to the detection of C. trachomatis infection in urine specimens collected from male and female subjects (VI). The differentiation between low-risk genotypes of human papilloma virus (HPV 6 or 11) from genotypes of high... 

In situ hybridization may be defined as the detection of nucleic acids in situ in cells, tissues, chromosomes, and isolated cell organelles. The technique was described in 1969 by two separate groups who demonstrated repetitive riboso-mal sequences in nuclei of Xenopus oocytes using radiolabeled probes (1,2). Refinements in recombinant DNA technology and the development of nonisotopic probe labeling and detection (3) obviate the need for radiation protection and disposal facilities, and have converted nonisotopic in situ hybridization (NISH) from a purely research technique to one that can be used in routine laboratory testing. 

Biochemical and localisation studies of the Hi receptor were made feasible with the design of reversible and irreversible radiolabelled probes such as [3H]mepyramine, [125l]iodobolpyramine and [125l]iodoazidophenpyramine [4-6]. 

Hybridize nitrocellulose membrane with radiolabeled probe. 

Release the DNA from the bacterial cells or phage using alkali. Neutralize and hybridize with radiolabeled probe. 

PBPs). A group at Lilly has introduced the radiolabeled probe 56 and fluorescence-labeled probe 57 as tool compounds to profile the PBPs in different bacterial strains using SDS-PAGE (Fig. 26) [100, 101]. At this time, no MS analysis of the protein bands has been performed. 

It is sometimes necessary to remove rapidly renaturing sequences from DNA samples before measuring the extent of relatedness, and this can be done on HA columns as described by Price et al.6 Self-annealing of the radiolabeled probe DNA can be minimized if the ratio of target to probe is kept to at least 1000 1. It may also be useful to determine the amount of self-annealing to be certain that this does not bias the calculated relatedness. Finally, a simplified version of the HA procedure was described by Lachance24 in which HA-containing minicolumns maintained under controlled temperatures are used to separate the duplexed DNAs. 

Can identify the DNA fragment containing the sequence complementary to the radiolabeled probe... 

Printed membranes were hybridized with a purified 600 bp GAPDH insert and labeled with radioactive probes. Visible signal was detected after 5 min of exposure with a digoxigenated probe followed by 30 min incubation with the radiolabeled probe. Mock hybridization experiments verified that there was no observed, unspecific signal, confirming that inkjet printing using standard desktop printers can be used to directly transfer DNA onto membranes for further reactivity. 

---