| Autor: |
Yutaka Yaida, Thaddeus S. Nowak, Katsumi Yufu |
| Rok vydání: |
2000 |
| Předmět: |
|
| DOI: |
10.1016/s0076-6879(99)14107-7 |
| Popis: |
Publisher Summary Localization of administered agents within a tissue is a critical component of in vivo antisense studies, particularly in heterogeneous tissues such as brain. Oligonucleotides prelabeled in various ways have been used to monitor tissue distribution with reasonable success. A potential problem with this approach is the sustained presence of the labeling moiety beyond the lifetime of the parent molecule, resulting in dissociations between the distribution of intact oligonucleotide and that of cleaved label. An alternative approach is the use of in situ hybridization with a labeled complementary sequence to detect the administered oligonucleotide. Conditions are easily chosen that require essentially a full-length sequence to be present to form stable hybrids, thereby allowing this method to specifically detect intact oligonucleotide. Procedures are described in the chapter for the use of both radiolabeled and fluorescein-labeled probes to detect antisense oligonucleotides in sections of fresh-frozen and perfusion-fixed brain. While the methods described in this chapter address the detection of phosphorothioate (PS) and phosphodiester (PO) oligonucleotides, the general approach should be readily applicable to other constructs. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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