In situ hybridization technique to localize rRNA and mRNA in XXXX neurons.

@entity1 @entity398 @entity35 @entity1604 @entity30555 @entity3488
In situ hybridization provides a method for identifying cells that contain specific nucleic acid sequences. This report outlines an in situ hybridization procedure for @entity1 neural tissue. The method maintains morphological quality and produces excellent specificity. Seven tritiated nucleic acid probes were examined: two ribosomal RNA probes, a control pBR322 plasmid probe, two probes encoding portions of the gene for @entity398 , one probe each encoding a portion of @entity3488 glycoprotein, and neurophysin. Using cryostat-cut @entity35 brain sections, rRNA probes labeled the cytoplasm of all cells and the nucleoli of larger neurons. The plasmid probe failed to produce a strong signal. @entity398 and @entity3488 probes appropriately labeled the cytoplasm of hypothalamic magnocellular neurons. @entity3488 parvocellular neurons were not identified by the current method, and the shorter length neurophysin probe failed to produce a signal. Methodological variables were examined by counting autoradiographic grains in cells. The longer @entity398 probe produced a stronger signal than the shorter @entity398 and @entity3488 probes, and higher probe concentrations resulted in stronger signal. Hybridization could be abolished by tissue pretreatment with @entity30555 , and longer exposure time increased signal strength. The outlined fixation steps with fresh-frozen tissue produced a superior signal compared to @entity1604 -perfused tissue.
